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  • Inorganic Chemistry  (83,665)
  • Biochemistry and Biotechnology  (16,218)
  • transformation
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  • 1
    Electronic Resource
    Electronic Resource
    The changing geography of morbidity and mortality in post-communist Poland (2000)
    Springer
    GeoJournal 50 (2000), S. 97-100 
    Details
    ISSN: 1572-9893
    Keywords: disease ; health ; morbidity ; mortality ; pollution ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract In general, the health of Poles improved markedly in the thirty years after the Second World War, but there was some deterioration after 1989 before improvement resumed. Only in the case of cancer is there an upward trend and so Poles are now healthier than they have been at any time in the past. However there are sharp regional variations well exemplified by the incidence of tuberculosis, where there appears to be some correlation with poorer housing and atmospheric pollution. High death rates in Lodz (consistently the highest in the country at the voivodship level between 1989 and 1996), may also be linked with environmental pollution as well as the ageing of the population. Variations between town and country are small, but Poland shows up in a poor light when compared with other European countries. These are important issues for the administration and financing of the welfare services.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007127431508
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  • 2
    Electronic Resource
    Electronic Resource
    A computational procedure for joining separate map sheets (2000)
    Springer
    GeoJournal 52 (2000), S. 253-262 
    Details
    ISSN: 1572-9893
    Keywords: map sheets ; GIS ; seamless spatial database ; transformation ; computational procedure
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract Map sheets have been often used as a basic spatial unit for managing spatial data produced from paper maps. This often results in incompatibility between adjacent map sheets, because spatial objects do not cross the boundaries smoothly and even the boundaries themselves do not match their neighbors exactly. To solve the problem this paper proposes a computational procedure for joining separate map sheets to obtain seamless spatial data. Line objects digitized separately in different map sheets are considered, which are frequently used to represent road networks, gas pipelines, and boundaries of polygon objects. The procedure consists of three steps: (1) extraction of end nodes, (2) detection of matching nodes, and (3) transformation of the map sheet. Each step goes interactively so that unexpected errors can be avoided by human observation. To test the validity of the procedure, map sheets are combined containing the road network data of Tokyo 23-ku area, Japan.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1014224325346
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  • 3
    Electronic Resource
    Electronic Resource
    Socioeconomic transformation in rural Romania through the eyes of experts: Demographic and social issues (2000)
    Springer
    GeoJournal 50 (2000), S. 151-155 
    Details
    ISSN: 1572-9893
    Keywords: demography ; experts ; rural ; Romania ; rural development ; social issues ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract There is no standard model of transformation for post- socialist countries and each country encounters specific problems rooted in the geographical characteristics of the areas concerned. The human resources are of the greatest importance because it really matters how people (especially the decision-makers) perceive system change and continually reformulate their expectations and strategies; so investigations into the views of people caught up in the transformation can provide a deeper understanding of the background to structural change. Working the national, regional level and local levels in Romania, experts were asked to consider the advantages and disadvantages arising out of the transformation, the most important problems and constraints for future rural development and the policies needed. The paper examines the responses on demographic and social issues. It emerges the most detailed responses were supplied by local-level representatives while respondents at the regional level steered a middle course between the need to address local problems and the prime importance of stimulating the Romanian economy so as to generate resources for welfare programmes (with the latter issue the overriding concern of interviewees at national level). There was general agreement on the importance of foreign investment and European integration for economic development, with local actors taking only small steps in line with the existing opportunities.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007120608735
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  • 4
    Electronic Resource
    Electronic Resource
    Transformation of batridene (2000)
    Springer
    Chemistry of natural compounds 36 (2000), S. 137-139 
    Details
    ISSN: 1573-8388
    Keywords: 1,1′,6,6′,7,7′-hexahydroxy-3,3′-dimethyl-5,5′-diisopropyl-2,2′-dinaphthylidene-8,8′-dibarbituric acid ; transformation ; DMSO
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Transformations of 1,1′,6,6′,7,7′-hexahydroxy-3,3′-dimethyl-5,5′-diisopropyl-2,2′-dinaphthylidene-8,8′-dibarbituric acid (batridene) in DMSO are studied.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02236415
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  • 5
    Electronic Resource
    Electronic Resource
    Metal resistance in Pseudomonas strains isolated from soil treated with industrial wastewater (2000)
    Springer
    World journal of microbiology and biotechnology 16 (2000), S. 177-182 
    Details
    ISSN: 1573-0972
    Keywords: Conjugation ; metal resistance ; plasmid DNA ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Heavy metal concentrations in soil treated with industrial wastewater of Aligarh City (U.P.), India were determined. The analysis of test samples revealed high levels of Fe, Zn, Ni and Cu. A total of 45 Pseudomonas spp. were isolated from soil and were characterized on the basis of morphological, cultural and biochemical characteristics. MICs of Hg2+, Cd2+, Cu2+, Cr3+, and Zn2+ for each isolate were determined. Eighty percent of the strains isolated from soil harboured resistance to copper, whereas 73.3% of the isolates exhibited resistance to cadmium, 71.1% to chromium and zinc and 48.8% to mercury. A maximum MIC of 200 μg/ml for mercury and 1600 μg/ml for other metals was observed. Metal resistance was found to be plasmid mediated as evidenced by transformation studies. Further, the transmissible nature of chromium resistance was confirmed by conjugation. Agarose gel electrophoresis using the miniprep method for plasmid isolation revealed that these isolates harboured plasmids of molecular weights (45 & 47 kb) using EcoRI and HindIII digests of λDNA and undigested λDNA as standard markers.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008905902282
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  • 6
    Electronic Resource
    Electronic Resource
    The effect of DNA/gold particle preparation technique, and particle bombardment device, on the transformation of barley (Hordeum vulgare) (2000)
    Springer
    Euphytica 111 (2000), S. 67-76 
    Details
    ISSN: 1573-5060
    Keywords: Barley ; Hordeum vulgare ; transformation ; particle bombardment ; particle gun
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Immature embryos of the spring barley variety GoldenPromise, were bombarded with three different particledelivery systems and both transient and stabletransformation examined. In addition, a range oftechniques for the preparation of the DNA coated goldparticles was examined. Fertile transgenic barleyplants were obtained using three particle preparationtechniques which differed in the amount of gold andDNA used for each bombardment. However, only one ofthe particle delivery systems, the PDS 1000/He device,appeared to be effective in yielding transformedbarley plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1003700300235
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  • 7
    Electronic Resource
    Electronic Resource
    Towards genetic transformation in the monocot Alstroemeria L. (2000)
    Springer
    Euphytica 115 (2000), S. 17-26 
    Details
    ISSN: 1573-5060
    Keywords: cell suspension ; monocotyledon ; selection ; somatic embryogenesis ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The successful application of plant biotechnology to Alstroemeria improvement will largely depend on the availability of an efficient regeneration/transformation system. Regeneration in Alstroemeria is accomplished from nodular embryogenic callus initiated from zygotic embryos. Histological studies of embryogenic callus initiation from 4-weeks old cultured ovules revealed that the outermost layers of the protoderm of the embryogenic nodules divided to form either a new nodule or aproembryo. Transient gene expression after particle bombardment of nodular embryogenic callus was optimized using DNA of pAHC25. The highest β-glucuronidase expression was found when the GUS gene was under control of the maize ubiquitin promoter, the target tissue was placed 5 cm below the microcarrier launch assembly and when the rupture disc-breakage point was between 650–900 psi. Kanamycin blocked regeneration of somatic embryos, however, did not block growth of nodular embryogenic callus. With phosphinothricin both callus growth and regeneration were blocked. Bombardment of nodular embryogenic callus with DNA of pAHC25 combined with selection on medium containing phosphinothricin resulted in putative transgenic chimeric. Friable calli were selected from nodular embryogenic callus and used to initiate suspensions. These cell suspensions were subjected to transformation by particle bombardment using DNA of pAHC25 and resulted in a stable transformed friable callus line after selection based on luciferase activity. Even after 2 years of maintenance this callus line was luciferase positive and the Polymerase Chain Reaction analysis demonstrated the presence of the introduced gene in this friable callus line.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1003979423469
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  • 8
    Electronic Resource
    Electronic Resource
    Rapid transformation and regeneration of alfalfa (Medicago falcata L.) via direct somatic embryogenesis (2000)
    Springer
    Plant growth regulation 31 (2000), S. 155-166 
    Details
    ISSN: 1573-5087
    Keywords: alfalfa ; cell division cycle ; somatic embryogenesis ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Two simple, rapid and efficient protocols for theregeneration of transformed tetraploid lines ofalfalfa (Medicago falcata L.) have beendeveloped and compared. Leaf explants fromembryogenic lines 47/1-150 and 47/1-5 were inoculatedwith Agrobacterium tumefaciens containingconstructs carrying the nptII selectable markergene and promoter:gusA gene fusions under thecontrol of the CaMV 35S or Arabidopsis cdc2a,CycB1 and CycA2 promoters. In the firstregeneration system (the MSH system), inoculated leafexplants were incubated on MS medium supplemented with2,4-D and kinetin and then subcultured onto plantgrowth regulator-free MS medium in order to inducedirect somatic embryogenesis. In the secondregeneration system (the B5h system), the inoculatedexplants were incubated on B5h medium to induceindirect production of somatic embryos viaembryogenic callus. In both systems, an effectivekanamycin selection regime was employed and wasmaintained when the embryos were subcultured onto arecovery medium (Boi2Y) to promote further embryodevelopment. The use of Boi2Y medium was particularlyimportant for shortening the regeneration time andpromoting a higher frequency of healthy plantletproduction from the somatic embryos. The maturesomatic embryos were finally transferred to plantgrowth regulator-free MS medium for plantletformation. Transgenic plantlets were produced within10–14 weeks in the MSH system and 12–16 weeks in theB5h system. The MSH system appears to be the fastesttransformation system reported for leguminous speciesto date. Confirmation of transformation was obtainedusing a re-callusing assay on kanamycin and subsequentSouthern blot hybridisation and PCR analysis. Theability to induce expression of GUS activity in leafexplants containing the cell division cycle genepromoter:gusA constructs by 2,4-D treatment alsoproved to be a reliable indicator of transformation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006306909722
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  • 9
    Electronic Resource
    Electronic Resource
    The Effects of Molybdenum on the High-Temperature Oxidation Resistance of Thin Foils of Fe–20Cr–5Al at Very High Temperatures (2000)
    Springer
    Oxidation of metals 53 (2000), S. 561-576 
    Details
    ISSN: 1573-4889
    Keywords: Fe–Cr–Al alloy ; oxidation ; molybdenum ; breakaway oxidation ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics
    Notes: Abstract Thin foils of Fe–20Cr–5Al alloys are susceptible to breakawayoxidation once the aluminum content of the substrate has fallen below somecritical value. The combined addition of 0.1 wt.% lanthanum and 0, 1, or 2wt% molybdenum has a beneficial effect on the high-temperature oxidation ofsuch foils. Lanthanum has the well-known reactive-element effect on adhesionof the protective alumina scale, thereby increasing the time to onset ofbreakaway oxidation, while, for alloys containing molybdenum, breakawayoxide spreads relatively slowly over the specimen in comparison to alloysthat contain no molybdenum. In particular, molybdenum-containing alloys areable to develop a protective Cr2O3 layer at the breakawayoxide–substrate interface. Conversely, molybdenum-free alloys form aninternal-oxide zone in the substrate adjacent to this interface, rather thana Cr2O3 layer, so breakaway oxide spreads rapidly. A martensitic phase isobserved in the substrate adjacent to the breakaway oxide formed on Fe–20Cr–5Al–La specimens, which means that theα-phase has transferred to the γ -phase at the temperature ofthe oxidation test (1150°C). Conversely, α-phase is retained inthe molybdenum-containing alloy, even after breakaway takes place, sincemolybdenum, which is a strong ferrite former, is enriched in the alloyadjacent to areas of breakaway oxide. The diffusion rate of chromium isslower in the γ than in the α-phase so a continuouschromium-rich oxide layer, which is effective in inhibiting breakawayoxide from spreading, cannot be established at the breakawayoxide–substrate interface for the molybdenum-free alloys.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1004689211302
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  • 10
    Electronic Resource
    Electronic Resource
    Factors involved in Agrobacterium tumefaciens-mediated gene transfer into Pinus nigra Arn. ssp. salzmannii (Dunal) Franco (2000)
    Springer
    Euphytica 114 (2000), S. 195-203 
    Details
    ISSN: 1573-5060
    Keywords: conifers ; salgareño pine ; tissue culture ; transformation ; transient gene expression ; uidA expression ; vir gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Cotyledons from dissected sterile embryos of salgareño pine (Pinus nigra Arn. ssp. salzmannii (Dunal) Franco) were inoculated with different disarmed Agrobacterium tumefaciens strains harbouring the binary vector p35SGUSint. The transient expression of a β-glucuronidase gene (uidA) was studied, using a histochemical staining procedure. Nineteen days after inoculation, the activity of β-glucuronidase was detected in epidermal and subepidermal layers of cotyledonary explants. The EHA105 strain harbouring a disarmed agropine-type Ti-plasmid (pTiBO542) was the most effective for gene transfer of the uidA gene. The effects of exudates and extracts from 0-day-old embryos on induction of vir gene expression in A. tumefaciens were also examined. The results of this study showed that salgarño pine embryo exudates contain a substance(s) that induce vir gene expression, in similar way to that observed with 100 μM acetosyringone (AS).All these findings suggest that T-DNA processing and transfer might take place when Agrobacterium infects suitable tissues of salgareño pine.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1003946131356
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  • 11
    Electronic Resource
    Electronic Resource
    Metamorphosis of a Non-Functioning Pituitary Adenoma to Cushing's Disease (2000)
    Springer
    Pituitary 3 (2000), S. 117-122 
    Details
    ISSN: 1573-7403
    Keywords: Cushing's disease ; silent ; pituitary ; tumour ; macroadenoma ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Silent pituitary adenomas occur in 25–40% of all clinically apparent pituitary tumours. However, the subsequent development of florid Cushing's disease in a patient with a previous non-functioning tumour is extremely rare. We describe a 47 year-old woman presenting initially with a large, invasive and recurrent, non-functioning pituitary tumour. Histopathologic study of the initial tissue did not stain for any hormones. Six years after the initial presentation, she manifested florid ACTH dependent Cushing's syndrome. A recurrent invasive pituitary macroadenoma necessitated a third transphenoidal surgery. The resected specimen, in this instance, revealed positive staining for ACTH, FSH, prolactin, and growth hormone on immunocytochemistry. An incomplete response to surgical, radiation and medical therapy necessitated a bilateral adrenalectomy to control the hypercortisolism, which resulted in remarkable clinical improvement. We also review five previous case reports from the revision literature of similar transformations from non-secreting pituitary adenomas to Cushing's disease. This subset of patients may represent yet another entity in the widening spectrum of Cushing's syndrome.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009961925780
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  • 12
    Electronic Resource
    Electronic Resource
    Floral Spray Transformation Can Efficiently Generate Arabidopsis (2000)
    Springer
    Transgenic research 9 (2000), S. 471-486 
    Details
    ISSN: 1573-9368
    Keywords: Arabidopsis thaliana ; Agrobacterium tumefaciens ; floral spray ; SOD ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In this study, floral spray and floral dip were used to replace the vacuum step in the Agrobacterium-mediated transformation of a superoxide dismutase (SOD) gene into Arabidopsis. The transgene was constructed by using a CaMV 35S promoter to drive a rice cytosolic CuZnSOD coding sequence in Arabidopsis. The transgene construct was developed in binary vectors and mobilized into Agrobacterium. When Arabidopsis plants started to initiate flower buds, the primary inflorescence shoots were removed and then transformed by floral spray or floral dip. More than 300 transgenic plants were generated to assess the feasibility of floral spray used in the in planta transformation. The result indicates that the floral spray method of Agrobacterium can achieve rates of in planta transformation comparable to the vacuum-infiltration and floral dip methods. The floral spray method opens up the possibility of in planta transformation of plant species which are too large for dipping or vacuum infiltration.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026522104478
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  • 13
    Electronic Resource
    Electronic Resource
    Efficient Production of Transgenic Cassava Using Negative and Positive Selection (2000)
    Springer
    Transgenic research 9 (2000), S. 405-415 
    Details
    ISSN: 1573-9368
    Keywords: Manihot esculenta ; transformation ; Agrobacterium tumefaciens ; mannose ; hygromycin ; selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to improve the efficiency of cassava (Manihot esculenta Crantz) transformation, two different selection systems were assessed, a positive one based on the use of mannose as the selective agent, and a negative one based on hygromycin resistance encoded by an intron-containing hph gene. Transgenic plants selected on mannose or hygromycin were regenerated for the first time from embryogenic suspensions cocultivated with Agrobacterium. After the initial selection using mannose and hygromycin, 82.6% and 100% of the respective developing embryogenic callus lines were transgenic. A system allowing plant regeneration from only transgenic lines was designed by combining chemical selection with histochemical GUS assays. In total, 12 morphologically normal transgenic plant lines were produced, five using mannose and seven using hygromycin. The stable integration of the transgenes into the nuclear genome was verified using PCR and Southern analysis. RT-PCR and northern analyses confirmed the transgene expression in the regenerated plants. A rooting test on mannose containing medium was developed as an alternative to GUS assays in order to eliminate escapes from the positive selection system. Our results show that transgenic cassava plants can be obtained by using either antibiotic resistance genes that are not expressed in the micro-organisms or an antibiotic-free positive selection system.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026509017142
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  • 14
    Electronic Resource
    Electronic Resource
    Two types of new algorithms for finding explicit analytical solutions of nonlinear differential equations (2000)
    Springer
    Applied mathematics and mechanics 21 (2000), S. 1423-1431 
    Details
    ISSN: 1573-2754
    Keywords: nonlinear differential equations ; transformation ; algorithm ; analytical solution ; O175.29
    Source: Springer Online Journal Archives 1860-2000
    Topics: Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics , Mathematics , Physics
    Notes: Abstract The idea of AC=BD was applied to solve the nonlinear differential equations. Suppose that Au=0 is a given equation to be solved and Dv=0 is an equation to be easily solved. If the transformation u=Cv is obtained so that v satisfies Dv=0, then the solutions for Au=0 can be found. In order to illustrate this approach, several examples about the transformation C are given.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02459221
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  • 15
    Electronic Resource
    Electronic Resource
    Transgenic Trifolium repens with foliage accumulating the high sulphur protein, sunflower seed albumin (2000)
    Springer
    Transgenic research 9 (2000), S. 103-113 
    Details
    ISSN: 1573-9368
    Keywords: inheritance ; Rubisco small subunit promoters ; sonication ; sulphur nutrition ; sunflower albumin ; transformation ; white clover
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract With the aim of increasing the rumen-protected level of the sulphur amino acids cysteine and methionine in Trifolium repens, we introduced the coding sequence of the sunflower seed albumin (SSA) into T. repens by Agrobacterium tumefaciens-mediated transformation. The SSA gene was modified such that the protein would be localised to the endoplasmic reticulum (ER). Four different T-DNA constructions all containing the SSA gene driven by either the promoter of a gene encoding the small subunit of ribulose bisphosphate carboxylase (Rubisco) from Arabidopsis thaliana (A ssu), the promoter of the gene encoding the small subunit of Rubisco of Medicago sativa (L ssu), or the Cauliflower Mosaic Virus 35S promoter (CaMV35S), were transferred to T. repens cv. Haifa. Transgenic T 0-plants and inter-transgenic hybrids were analysed for the level of SSA accumulation in the leaves by western blotting. The highest observed level of SSA accumulation was 0.1% of total extractable leaf protein. We observed that the promoter had a substantive effect on the level of SSA accumulation with A ssu〉CaMV35S〉L ssu. Results from the inter-transgenic hybrids showed that the capacity to synthesise SSA was inherited. However the level of SSA accumulation in the leaves generally appears not to be additive with extra transgenic loci. During this work, we attempted to improve the efficiency of A. tumefaciens-mediated transformation of T. repens using the SAAT-method (Sonication Assisted Agrobacterium-mediated Transformation) on cotyledons of T. repens. T-DNA transfer was in general not enhanced by sonication compared to traditional A. tumefaciens-mediated transformation. Furthermore, Southern blot analyses of plants regenerated from the same cotyledon after A. tumefaciens treatment and under selection, indicated that multiple shoots were usually derived from the same transformation event. We concluded from these results that only one plant from each A. tumefaciens-treated cotyledon should be taken to avoid transgenic clones.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008967409302
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  • 16
    Electronic Resource
    Electronic Resource
    Transformation of the tropane alkaloid-producing medicinal plant Hyoscyamus muticus by particle bombardment (2000)
    Springer
    Transgenic research 9 (2000), S. 163-168 
    Details
    ISSN: 1573-9368
    Keywords: Hyoscyamus muticus ; particle bombardment ; transformation ; tropane alkaloids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report an efficient whole plant transformation system for Hyoscyamus muticus, an important medicinal plant of the Solanaceous family. We developed a system using a plasmid carrying the nptII and gusA genes, which was delivered into leaf explants by particle bombardment. Ten percent of bombarded leaf explants formed kanamycin-resistant callus, from which putative transgenic plants were recovered. The nptII gene conferring kanamycin resistance was found to be incorporated into the genome of all transgenic plants screened. Over 50% of the kanamycin resistant plants showed strong expression of the non-selected gusA gene. The majority of transgenic plants reached maturity, could be self pollinated, and produced fertile seed. A simple and efficient whole plant transformation system for this medicinal plant is an important step in furthering our understanding of tropane alkaloid production in plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008912330067
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  • 17
    Electronic Resource
    Electronic Resource
    Enhanced forskolin production in genetically transformed cultures of Coleus forskohlii by casein hydrolysate and studies on growth and organisation (2000)
    Springer
    Biotechnology letters 22 (2000), S. 133-136 
    Details
    ISSN: 1573-6776
    Keywords: casein hydrolysate ; Coleus forskohlii ; forskolin ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Casein hydrolysate at 2.0 g l−1 significantly enhanced forskolin content (2.3 mg g−1 cell dry wt) in a rhizogenic tumourous line, GCO-RCH-2 of Coleus forskohlii. In rooty teratoma line, RC-ST-2/4, forskolin content enhanced to 1.7 mg g−1 cell dry wt in presence of 2.5 g l−1 casein hydrolysate. Unlike untransformed calli and rhizogenic/root cultures, all the forskolin yielding transformed cultures of C. forskohlii have been maintained for over 5 years.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005618307074
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  • 18
    Electronic Resource
    Electronic Resource
    Agrobacterium-mediated sorghum transformation (2000)
    Springer
    Plant molecular biology 44 (2000), S. 789-798 
    Details
    ISSN: 1573-5028
    Keywords: Agrobacterium tumefaciens ; sorghum ; transformation ; transgenic plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Agrobacterium tumefaciens was used to genetically transform sorghum. Immature embryos of a public (P898012) and a commercial line (PHI391) of sorghum were used as the target explants. The Agrobacterium strain used was LBA4404 carrying a `Super-binary' vector with a bar gene as a selectable marker for herbicide resistance in the plant cells. A series of parameter tests was used to establish a baseline for conditions to be used in stable transformation experiments. A number of different transformation conditions were tested and a total of 131 stably transformed events were produced from 6175 embryos in these two sorghum lines. Statistical analysis showed that the source of the embryos had a very significant impact on transformation efficiency, with field-grown embryos producing a higher transformation frequency than greenhouse-grown embryos. Southern blot analysis of DNA from leaf tissues of T0 plants confirmed the integration of the T-DNA into the sorghum genome. Mendelian segregation in the T1 generation was confirmed by herbicide resistance screening. This is the first report of successful use of Agrobacterium for production of stably transformed sorghum plants. The Agrobacterium method we used yields a higher frequency of stable transformation that other methods reported previously.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026507517182
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  • 19
    Electronic Resource
    Electronic Resource
    Bringing “The Dead” to Life: Reading James Joyce (2000)
    Springer
    Journal of applied psychoanalytic studies 2 (2000), S. 109-115 
    Details
    ISSN: 1573-3459
    Keywords: James Joyce ; “The Dead” ; fiction ; reading ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Psychology
    Notes: Abstract The author offers a personal reading of James Joyce's “The Dead.” She focuses on how the sounds of the language are used to portray the main character's deadness and his beginning to come to life once the barriers to inner self and self-knowledge are broken.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1010193724350
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  • 20
    Electronic Resource
    Electronic Resource
    Changes in lipid and protein constituents of rafts and caveolae in multidrug resistant cancer cells and their functional consequences (2000)
    Springer
    Glycoconjugate journal 17 (2000), S. 253-259 
    Details
    ISSN: 1573-4986
    Keywords: Cancer ; caveolae ; caveolin ; cholesterol ; glucosylceramide ; multidrug resistance ; rafts ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract The carcinogenic process involves a complex series of genetic and biochemical changes that enables transformed cells to proliferate, migrate to secondary sites and, in some cases, acquire mechanisms that make cancer cells resistant to chemotherapy. This phenomenon in its most common form is known as multidrug resistance (MDR). It is usually mediated by overexpression of P-glycoprotein (P-gp) or other plasma membrane ATPases that export cytotoxic drugs used in chemotherapy, thereby reducing their efficacy. However, additional adaptive changes are likely to be required in order to confer a full MDR phenotype. Recent studies have shown that acquisition of MDR is accompanied by up-regulation of lipids and proteins that constitute lipid rafts and caveolar membranes, notably glucosylceramide and caveolin. These changes may be related to the fact that in MDR cells a significant fraction of cellular P-gp is associated with caveolin-rich membrane domains, they may be involved in drug transport and they could have an impact on drug-induced apoptosis and on the phenotypic transformation of MDR cancer cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026553626537
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  • 21
    Electronic Resource
    Electronic Resource
    Study of the factors influencing Agrobacterium-mediated transformation of pea (Pisum sativum L.) (2000)
    Springer
    Molecular breeding 6 (2000), S. 185-194 
    Details
    ISSN: 1572-9788
    Keywords: Agrobacterium ; transformation ; pea ; Pisum sativum L. ; PCR analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Factors influencing the efficiency of Agrobacterium-mediated transformation of pea were tested using highly efficient, direct regeneration system. The virulence of three Agrobacterium strains (octopine LBA 4404, nopaline C58C1 and succinamopine, hypervirulent EHA 105) clearly varied giving 1 transgenic plant per 100 explants for LBA 4404, 2.2 for C58C1 and 8.2 for EHA 105. To test the efficacy of selection agents we used the hypervirulent EHA 105 strain carrying pGPTV binary vector with one of four different selection genes: nptII, hpt, dhfr or bar. The mean number of transgenic, kanamycin-resistant plants for two cultivars tested was 4.2 per 100 explants and was slightly higher than the number of phosphinothricin-resistant plants (3.6 plants per 100 explants). The proportion of transgenics among kanamycin-selected plants was also higher than among phosphinothricin-resistant plants (35% and 28% respectively). There was no regeneration on hygromycin or methotrexate media (transformation with hpt and dhfr genes). Acetosyringone had no apparent influence on efficiency of transformation with hypervirulent EHA 105 strain, however it did affect the rate of transformation when moderately virulent C58C1 was used. Recovery of transgenic plants was enhanced after application of 5-azacytidine. The presence of integrated T-DNA was checked by PCR and confirmed by Southern hybridization. T-DNA was stably transmitted to the next generation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009679908948
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  • 22
    Electronic Resource
    Electronic Resource
    Field resistance to Tomato spotted wilt virus in transgenic peanut (Arachis hypogaea L.) expressing an antisense nucleocapsid gene sequence (2000)
    Springer
    Molecular breeding 6 (2000), S. 227-236 
    Details
    ISSN: 1572-9788
    Keywords: antisense DNA ; co-transformation ; nucleocapsid gene ; pathogen-derived resistance ; somatic embryogenesis ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Peanut (Arachis hypogaea L.) lines transgenic for the antisense nucleocapsid (N) gene of a Tomato spotted wilt virus (TSWV) strain isolated from peanut were generated by microprojectile-mediated transformation of repetitive somatic embryos of cultivars VC1 and AT120. The selectable marker (hygromycin resistance) and the N gene were on separate plasmids. A total of 207 VC1 and 120 AT120 hygromycin-resistant lines were produced. Of all the VC1 plants recovered 71% were cotransformed with the N gene (N+), but all plants were sterile. For AT120, 48 of the transgenic cell lines converted into plants. Polymerase chain reaction (PCR) screening showed 15 of the lines were transgenic for the N gene (N+), and two of these lines were fertile. A field test was conducted in 1998 at Ashburn, GA, using seeds from each fertile line, along with segregated and non-transgenic controls. Plants from four randomly selected field plots were examined for symptoms and analyzed by double-antibody sandwich enzyme-linked immunoabsorbent assay and PCR at 10 and 14 weeks after planting. At 14 weeks, 76% of the N+ plants were symptomless, while 2% were severely symptomatic or dead. In contrast, only 42% of the plants lacking the N gene were symptomless and 50% were severely symptomatic or dead. Northern blot analysis of selected field-resistant plants detected transgene RNA, and the transcript level appeared undiminished after viral exposure.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009649408157
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  • 23
    Electronic Resource
    Electronic Resource
    Insect- and herbicide-resistant transgenic eucalypts* (2000)
    Springer
    Molecular breeding 6 (2000), S. 307-315 
    Details
    ISSN: 1572-9788
    Keywords: bar ; cry3A ; Eucalyptus ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Transgenic Eucalyptus camaldulensis containing both the insecticidal cry3A gene and the bar gene (conferring tolerance to the herbicide glufosinate ammonium) have been produced by Agrobacterium tumefaciens-mediated transformation of seedling explants. Transgenic plants from two lines tested were resistant to first instars of chrysomelid beetles that are important pests of commercial Australian eucalypt plantations. Both lines also exhibit tolerance to the broad-spectrum herbicide Liberty® at 6 l/ha (1.2 kg active ingredient per hectare), twice the field application rate. Transgenic insect- and herbicide-resistant eucalypts like these are likely to provide better insect and weed control options in plantations, particularly during the vulnerable establishment phase, provided that any adverse ecological impacts of releasing transgenic trees into the environment can be assessed and minimized.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009676214328
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  • 24
    Electronic Resource
    Electronic Resource
    Transformation of Japanese persimmon (Diospyros kaki Thunb.) with a bacterial gene for choline oxidase (2000)
    Springer
    Molecular breeding 6 (2000), S. 501-510 
    Details
    ISSN: 1572-9788
    Keywords: choline oxidase ; glycinebetaine ; Japanese persimmon ; salt tolerance ; transformation ; woody plant
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract This report describes the first successful genetic engineering of tolerance to salt in an agriculturally important species of woody plants by Agrobacterium-mediated transformation with the codA gene of Arthrobacter globiformis. This gene encodes choline oxidase, which catalyzes the oxidation of choline to glycinebetaine. The binary plasmid vector pGC95.091, containing a kanamycin-resistance gene (nptII), a gene for β-glucuronidase (gusA) and the codA gene in its T-DNA region, was used with a disarmed strain of Agrobacterium tumefaciens, EHA101, to transform Japanese persimmon (Diospyros kaki Thunb. `Jiro') by the leaf disk transformation method. The pRS95.101 plasmid that included only nptII and gusA in the T-DNA region was used as a control. We selected eight transgenic lines with one or two copies of the T-DNA after transformation with pGC95.091 (PC lines) and three lines after transformation with pRS95.101 (PR lines). The eight PC lines produced choline oxidase and glycinebetaine whereas neither was found in untransformed `Jiro' and in the control PR lines. Transgenic plants grew normally, resembling wild-type plants both in vitro and ex vitro. The activity of photosystem II in leaves of the transgenic Japanese persimmon plants under NaCl stress was determined in terms of the ratio of the variable (F v) to the maximum (F m) fluorescence of chlorophyll (F v/F m). The rate of decline in (F v/F m under NaCl stress was lower in the PC lines than in the control PR lines. These results demonstrated that genetic engineering of Japanese persimmon, which allowed it to accumulate glycinebetaine, enhanced the tolerance to salt stress of this plant.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026513831290
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  • 25
    Electronic Resource
    Electronic Resource
    Generation of transgenic wheat (Triticum aestivum L.) for constitutive accumulation of an Aspergillus phytase (2000)
    Springer
    Molecular breeding 6 (2000), S. 195-206 
    Details
    ISSN: 1572-9788
    Keywords: heterologous protein accumulation ; phytate phosphorus digestibility ; phytase ; phytic acid ; transformation ; Triticum aestivum L.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The Aspergillus niger phytase-encoding gene (phyA) has been constitutively expressed in wheat. Transgenic wheat lines were generated by microprojectile bombardment of immature embryos, using the bar-Bialaphos selection system. The bar and the phyA gene expression were controlled by the maize ubiquitin-1 promoter. To ensure secretion and glycosylation of the microbial phytase, an expression cassette was designed (Ubi-SP-Phy) where an α-amylase signal peptide sequence was inserted between the promoter and the phytase coding region. A similar cassette was constructed without the signal peptide sequence (Ubi-Phy). Five lines of fertile wheat transformed with the Ubi-SP-Phy were generated and two lines with the Ubi-Phy construct. The inheritance of the phyA gene was monitored through three generations. Western blotting of leaf and seed derived protein revealed the presence of an immunoreacting polypeptide of the size expected for the Aspergillus phytase. Up to 25 days after pollination, the heterologous phytase was exclusively present in the pericarp-seed coat-aleurone fraction. Thereafter, it accumulated in the endosperm in amounts exceeding that found in the seed coat and aleurone. The phyA mRNA and derived protein could at no stage be detected in the embryo. The Ubi-SP-Phy transgenic seeds exhibited up to 4-fold increase of phytase activity while up to 56% increase was found in Ubi-Phy plants. It is concluded that a functional Aspergillus phytase can be produced in significant amounts in wheat grains. This may be of relevance for improving the phytate-phosphorus digestibility when wheat grains are used for non-ruminant animal feed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009690730620
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  • 26
    Electronic Resource
    Electronic Resource
    Transformation of Brassica juncea (L.) Czern with bacterial codA gene enhances its tolerance to salt stress (2000)
    Springer
    Molecular breeding 6 (2000), S. 489-499 
    Details
    ISSN: 1572-9788
    Keywords: Choline oxidase ; glycinebetaine ; transformation ; Brassica juncea ; salt stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The codA gene for biosynthesis of glycinebetaine from Arthrobacter globiformis was used for transforming Brassica juncea cv. Pusa Jaikisan (which lack any means to synthesize glycinebetaine) through Agrobacterium mediated transformation. The stable insertion of the codA gene in the shoots obtained on medium with kanamycin and hygromycin was confirmed by PCR analysis of the nptII gene. Southern hybridization with a codA probe further demonstrated its successful integration. Immunoblot analysis revealed the presence of choline oxidase demonstrating that the bacterial codA gene had been successfully transcribed and translated. The seeds of transgenic lines showed enhanced capacity to germinate under salt stress as compared to that of the wild type. Further, the seedlings of transgenic plants that expressed codA gene showed significantly higher growth than that of the wild type under salt stress conditions. These results demonstrated that the introduction of a biosynthetic pathway for glycinebetaine into Brassica juncea significantly enhanced their salt tolerance. Hence, homozygous genotypes of selected transformed lines can be exploited for improving the salt tolerance of the desirable cultivars of Brassica juncea through breeding programmes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026542109965
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  • 27
    Electronic Resource
    Electronic Resource
    Transforming Self and Society: A "Critical Appreciation" Model (2000)
    Springer
    Systemic practice and action research 13 (2000), S. 475-501 
    Details
    ISSN: 1573-9295
    Keywords: reflection ; transformation ; self-society dynamics ; critical systems thinking ; systemic intervention ; critical self-reflection ; ideology-critique ; critical appreciation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Sociology
    Notes: Abstract This paper sets out to uncover some relationships between reflection, discourse and action. By challenging and synthesizing some polemical arguments concerning the creation, maintenance, and transformation of self and society, a model of self-society dynamics that operates through reflection, discourse, and action is developed. The model of self-society dynamics brings together aspects of self-reflection and ideology-critique (explored in the paper), which it is suggested are required for any intervention (transformative action) to be grounded in locally meaningful ways. By creating a dialog community in which self- and group assumptions can be subjected to validation through discourse, it is proposed that a dynamic balance between individual needs and broader societal aims may be achieved. If individuals can be open to such discourse (i.e., they can become critically self-reflective), then it is argued that possibilities for achieving sustainable change will be significantly enhanced.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009541430809
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  • 28
    Electronic Resource
    Electronic Resource
    Polymorphism in Polyamide of Dytek®-A and Dodecanedioic Acid (1999)
    Springer
    Journal of thermal analysis and calorimetry 56 (1999), S. 1133-1140 
    Details
    ISSN: 1572-8943
    Keywords: branched diamine ; melting ; polyamides ; polymorphism ; transformation ; WXRD
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Our X-ray work of Dytek®-A, 2-methyl-pentamethylenediamine, containing polyamides shows polymorphism, whereas the polyamides with linear diamines do not. The polyamide of Dytek®-A and dodecanedioic acid, MPMD-12, is singled out for discussion and compared with the unbranched analogs of polyamides 6,12 and 5,12. Due to the presence of the -CH3 side group in the 2-position of the diamine, the polyamide MPMD-12 exhibits two stable crystal conformations. The new δ polymorph is not seen in linear polyamides 6,12 and 5,12. Studies by DSC polyamide MPMD-12 clearly illustrates at least two crystal forms, γ and δ, coexisting over a wide temperature range, and the isolation of each phase is possible by controlling temperature and time. The DMA modulus in the temperature region between the glass transition (or alpha relaxation) and melting transition shows strong dependence on the thermal history as demonstrated in a study of crystallization kinetics.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1010105028316
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  • 29
    Electronic Resource
    Electronic Resource
    A Modular Vector for Agrobacterium Mediated Transformation of Wheat (1999)
    Springer
    Plant molecular biology reporter 17 (1999), S. 323-331 
    Details
    ISSN: 1572-9818
    Keywords: Agrobacterium ; modular vector ; transformation ; wheat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Wheat (cv Chinese Spring) tissues were transformed using Agrobacterium tumefasciens and a new plasmid modular vector, pMVTBP. We constructed pMVTBP with unique restriction sites connecting (1) the CaMV 35S promoter, (2) a Kozak sequence, (3) the FLAG epitope, (4) the (His)6 epitope, (5) a coding region (for wheat TATA Binding Protein, wTBP) and (6) the CaMV 35S 3′UTR. This vector thus allows easy exchange of different regulatory or coding sequences. Explants of either germinating mature seeds, or immature embryos, were induced to callus for up to two weeks, treated with virulence-induced bacteria for one hour, then regenerated into plantlets. Transient expression of a GUS reporter gene, assayed at about one week, occurred in 10–12% of calluses. Expression of the FLAG-tagged wTBP was also detected, by immunostaining. Stable expression, by selective growth on geneticin, and by GUS expression at about six weeks, occurred in 1–2% of calluses, quite comparable to that achieved by other methods.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007686408369
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  • 30
    Electronic Resource
    Electronic Resource
    Urban development in East Germany — specific features of urban transformation processes (1999)
    Springer
    GeoJournal 49 (1999), S. 43-51 
    Details
    ISSN: 1572-9893
    Keywords: housing market ; suburbanisation ; transformation ; urban development ; urban renewal ; East Germany
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract The aim of this paper is to analyse the main characteristics of post-socialist urban development in East Germany, especially the differences compared to urban development in other East and Central European countries. In spite of the many similar problems and processes in urban development, specific features of East Germany are characterised by the rapid growth of suburbia, especially in the first phase of transition, by the proceeding activities of urban renewal and revitalisation, and by a lower level of social polarisation and socio-spatial segregation as compared to other post-socialist countries. Important conditions for urban development in East Germany exist in special support programmes, high subsidies and other financial transfers as well as in engaged planning conceptions of the local authorities.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007016912703
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  • 31
    Electronic Resource
    Electronic Resource
    Wnt Signalling in Mammalian Development and Cancer (1999)
    Springer
    Cancer and metastasis reviews 18 (1999), S. 215-230 
    Details
    ISSN: 1573-7233
    Keywords: transformation ; tumour ; Frizzled ; Dishevelled ; glycogen synthase kinase-3β
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Wnt signalling is involved in a variety of mammalian developmental processes, including cell proliferation, differentiation and epithelial–mesenchymal interactions, through which they contribute to the development of tissues and organs such as the limbs, the brain, the reproductive tract and the kidney. Wnts are secreted ligands that control cell processes via at least two pathways, one of which, the ‘canonical’ Wnt signalling pathway, operates through the cytosolic stabilisation of a transcriptional co-factor, β-catenin. This is achieved by downregulating the activity of a β-catenin turnover complex. Evidence from tumour expression studies, transgenic animals and in vitro experiments suggests that inappropriate activation of the canonical Wnt signalling pathway is a major feature in human neoplasia and that oncogenic activation of this pathway can occur at many levels. Inappropriate expression of the Wnt ligand and Wnt binding proteins have been found in a variety of human tumours. Further downstream, dysregulation of the β-catenin turnover complex, by loss of the Adenomatous Polyposis Coli or Protein Phosphatase 2A proteins, or by activating mutations of β-catenin, has been found in several tumour types, and is believed to be a key step in neoplastic progression. Transcriptional targets of the Wnt pathway include the cellular oncogenes cyclin D1 and c-myc. Activation of the Wnt signalling pathway by various means can therefore be a primary cause in oncogenesis, affecting cell proliferation, morphology and contact inhibition, as well as co-operating with other oncogenes in multistep tumour progression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006369223282
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  • 32
    Electronic Resource
    Electronic Resource
    REMI (Restriction Enzyme Mediated Integration) and its Impact on the Isolation of Pathogenicity Genes in Fungi Attacking Plants (1999)
    Springer
    European journal of plant pathology 105 (1999), S. 221-229 
    Details
    ISSN: 1573-8469
    Keywords: mutagenesis ; transformation ; plant disease ; recombination ; plant pathogenic fungi
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Development of molecular techniques for phytopathogenic fungi aims at the identification of fungal genes whose products are essential for successful infection of the host plant. Initial approaches have relied on isolating candidate genes and generating null-mutations by homologous recombination. Unfortunately, the results of this strategy have not been overly successful. This has led to a search for alternatives which allow an unbiased identification of pathogenicity genes. One method, which has proved successful in several systems, is a tagging mutagenesis procedure termed restriction enzyme mediated integration (REMI). In this mini-review we describe this procedure and review its features and results of its use when applied to the identification of fungal genes required for disease development in planta.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008757414036
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  • 33
    Electronic Resource
    Electronic Resource
    Anaerobic transformation of 1,1,1-trichloroethane by municipal digester sludge (1999)
    Springer
    Biodegradation 10 (1999), S. 297-305 
    Details
    ISSN: 1572-9729
    Keywords: abiotic ; biological ; cell-free extract ; chloroethane ; dechlorination ; 1,1-dichloroethane ; 1,1-dichloroethene ; digester ; methanogenic ; transformation ; 1,1,1-trichloroethane
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Energy, Environment Protection, Nuclear Power Engineering , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Anaerobic transformations of 1,1,1-trichloroethane (TCA), 1,1-dichloroethane (DCA), and chloroethane (CA) were studied with sludge from a lab-scale, municipal wastewater sludge digester. TCA was biologically transformed to DCA and CA and further to ethane by reductive dechlorination. TCA was also converted to acetic acid and 1,1-dichloroethene (11DCE) by cell-free extract. 11DCE was further biologically converted to ethene. This pathway was confirmed by transformation tests of TCA, DCA and CA, by tests with cell-free extract, and by chloride release during TCA degradation. With cell-free extract, acetic acid accounted for approximately 90% of the TCA transformed; tests with live cells indicate that the fraction of TCA transformed by this pathway decreased with lower biomass. The dechlorination of DCA to CA and CA to ethane was not stoichiometric. A high rate of TCA removal was observed under the experimental conditions. The results indicate that removal of TCA in anaerobic digestion should be complete, but DCA and CA could persist in a normally operating digester.
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    URL: http://dx.doi.org/10.1023/A:1008336103968
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  • 34
    Electronic Resource
    Electronic Resource
    Biotransformation of cholesterol by Micrococcus species mediated by plasmid DNA (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 411-415 
    Details
    ISSN: 1573-0972
    Keywords: Electroporation ; Micrococcus species ; steroid biotransformation ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A steroid-biotransforming strain RJ6 was identified as Micrococcus roseus. This bacterium has a 10 kb plasmid pMQV10. Curing mediated through cultivation of the culture with a low concentration (200 ng/ml) of mitomycin C is described. Loss of cholesterol degradation (chol+) and streptomycin resistance (Smr) phenotypes as a consequence of the loss of plasmid indicate the extrachromosomal location of these two genes in this strain. An electroporation procedure was developed for transformation of cured strain of Micrococcus (RJC6) by plasmids. Frequency of greater than 105 transformants/μg DNA was achieved, which is 100-fold higher than the standard transformation procedure that yielded 5.3×103 transformants/μg DNA in the same strain.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008990910303
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  • 35
    Electronic Resource
    Electronic Resource
    Evolution of gene transfer in bacteria (1999)
    Springer
    World journal of microbiology and biotechnology 15 (1999), S. 1-6 
    Details
    ISSN: 1573-0972
    Keywords: Bacteria ; conjugation ; DNA ; evolution ; gene transfer ; transduction ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The transfer of genetic information by transformation, conjugation and transduction in bacteria occurs frequently in nature. These diverse gene transfer mechanisms in bacteria are the result of evolution and are not linked to reproduction as in eukaryotic organisms. In this review, gene transfer in bacteria will be considered from an evolutionary perspective.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008830914223
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  • 36
    Electronic Resource
    Electronic Resource
    Selectable marker-free transgenic plants without sexual crossing: transient expression of cre recombinase and use of a conditional lethal dominant gene (1999)
    Springer
    Plant molecular biology 40 (1999), S. 223-235 
    Details
    ISSN: 1573-5028
    Keywords: conditional lethal dominant gene ; Cre/loxP ; Nicotiana tabacum ; site-specific recombinase ; transformation ; transient expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Transgenic tobacco plants were produced that contained single-copy pART54 T-DNA, with a 35S-uidA gene linked to loxP-flanked kanamycin resistance (nptII) and cytosine deaminase (codA) genes. Retransformation of these plants with pCre1 (containing 35S transcribed cre recombinase and hygromycin (hpt) resistance genes) resulted in excision of the loxP-flanked genes from the genome. Phenotypes of progeny from selfed-retransformed plants confirmed nptII and codA excision and integration of the cre-linked hpt gene. To avoid integration of the hpt gene, and thereby generate plants totally free of marker genes, we attempted to transiently express the cre recombinase. Agrobacterium tumefaciens (pCre1) was cocultivated with leaf discs of two pART54-transformed lines and shoots were regenerated in the absence of hygromycin selection. Nineteen of 773 (0.25%) shoots showed tolerance to 5-fluorocytosine (5-fc) which is converted to the toxic 5-fluorouracil by cytosine deaminase. 5-fc tolerance in six shoots was found to be due to excision of the loxP-flanked region of the pART54 T-DNA. In four of these shoots excision could be attributed to cre expression from integrated pCre1 T-DNA, whereas in two shoots excision appeared to be a consequence of transient cre expression from pCre1 T-DNA molecules which had been transferred to the plant cells but not integrated into the genome. The absence of selectable marker genes was confirmed by the phenotype of the T1 progeny. Therefore, through transient cre expression, marker-free transgenic plants were produced without sexual crossing. This approach could be applicable to the elimination of marker genes from transgenic crops which must be vegetatively propagated to maintain their elite genotype.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006184221051
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  • 37
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    Electronic Resource
    Recovery of transgenic orchid plants with hygromycin selection by particle bombardment to protocorms (1999)
    Springer
    Plant cell, tissue and organ culture 58 (1999), S. 87-92 
    Details
    ISSN: 1573-5044
    Keywords: ß-glucuronidase ; dendrobium ; hygromycin phosphotransferase ; orchid ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protocorms of orchid (Dendrobium hybrid) were transformed by microprojectile bombardment with a helium-pressured PDS 1000 particle gun. Gold particles coated with plasmid DNA containing ß-glucuronidase (GUS) and hygromycin phosphotransferase (Hpt) marker genes were used. Potentially transformed tissues were identified by active growth on MS medium supplemented with 50mg l-1 hygromycin. After 4–6 months of continuous selection, 15 hygromycin-resistant lines were recovered. Integration of transgenes into the genome of the transformed protocorms and plantlets were confirmed by GUS histochemical assay and Southern blot hybridization. The transgenic protocorms have gone through propagation for more than 8 months and maintained their transgenic characters. These results indicate that we have established a system for orchid transformation in a relatively high frequency and the transgenes are stably expressed in the transgenic plants.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006334605947
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  • 38
    Electronic Resource
    Electronic Resource
    Isolation of state transition mutants of Chlamydomonas reinhardtii by fluorescence video imaging (1999)
    Springer
    Photosynthesis research 61 (1999), S. 43-51 
    Details
    ISSN: 1573-5079
    Keywords: Chlamydomonas reinhardtii ; LHC II phosphorylation ; mutagenesis ; Photosystem II redox control ; state 2 to state 1 transition ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Oxygenic photosynthetic organisms adapt to varying light conditions by changing the distribution of light energy between Photosystem II (PS II) and photosystem I (PS I) during so-called state transitions. To identify the genes involved in this process, we have exploited a simple chlorophyll fluorescence video-imaging technique to screen a library of nuclear mutants of Chlamydomonas reinhardtii for colonies grown on agar plates that are disturbed in their ability to regulate light energy distribution between PS I and PS II. Subsequent modulated fluorescence measurements at room temperature and 77 K fluorescence emission spectra confirmed that 5 mutants (0.025% of total number screened) were defective in state transitions. [32P]orthophosphate phosphorylation experiments in vivo revealed that in one of these mutants, designated stm1, the level of LHC II polypeptide phosphorylation was drastically reduced compared with wild type. Despite WT levels of PS I and PS II, stm1 grew photoautotrophically at reduced rates, compared with WT especially under low light conditions, which is consistent with an important physiological role for state transitions. Our results highlight the feasibility of video imaging in tandem with mutagenesis as a means of identifying the genes involved in controlling state transitions in eukaryotic photosynthetic organisms.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006229308606
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  • 39
    Electronic Resource
    Electronic Resource
    A mini binary vector series for plant transformation (1999)
    Springer
    Plant molecular biology 40 (1999), S. 711-717 
    Details
    ISSN: 1573-5028
    Keywords: Agrobacterium tumefaciens ; Arabidopsis thaliana ; binary vector ; T-DNA ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A streamlined mini binary vector was constructed that is less than 1/2 the size of the pBIN19 backbone (3.5 kb). This was accomplished by eliminating over 5 kb of non-T-DNA sequences from the pBIN19 vector. The vector still retains all the essential elements required for a binary vector. These include a RK2 replication origin, the nptIII gene conferring kanamycin resistance in bacteria, both the right and left T-DNA borders, and a multiple cloning site (MCS) in between the T-DNA borders to facilitate cloning. Due to the reduced size, more unique restriction sites are available in the MCS, thus allowing more versatile cloning. Since the traF region was not included, it is not possible to mobilize this binary vector into Agrobacterium by triparental mating. This problem can be easily resolved by direct transformation. The mini binary vector has been demonstrated to successfully transform Arabidopsis plants. Based on this mini binary vector, a series of binary vectors were constructed for plant transformation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006201910593
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  • 40
    Electronic Resource
    Electronic Resource
    Optimisation of procedures for microprojectile bombardment of microspore-derived embryos in wheat (1999)
    Springer
    Plant cell, tissue and organ culture 57 (1999), S. 207-210 
    Details
    ISSN: 1573-5044
    Keywords: biolistics ; gene expression ; haploid ; transformation ; Triticum aestivum
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Using the PDS-1000/He Biolistic® Particle Delivery System, the microprojectile travel distance, rupture disk pressure and DNA/gold particle concentrations were assessed in order to optimise short and longer-term β-glucuronidase reporter gene expression in microspore-derived embryos of wheat. The effects were also evaluated of using sterile filter paper to support explants and treatment with a high osmoticum medium (0.2 M mannitol/0.2 M sorbitol or 0.4 M maltose). In the optimised procedure, wheat microspore-derived embryos (MDEs), were placed on filter paper and incubated on medium containing 0.4 M maltose, for 4 h pre- and 45 h post-bombardment. Five μl pAHC25 (0.75 mg ml-1 in TE buffer) was precipitated onto 25 μl gold particles (60 mg ml-1 in sterile water), using 20 μl spermidine (0.1 M) and 50 μl CaCl2 (2.5 M). The particles were centrifuged and resuspended in 75 μl absolute ethanol prior to the preparation of 6 macrocarriers. A microprojectile travel distance of 70 mm, a rupture pressure of 1300 p.s.i., and a vacuum of 29′′ Hg were employed. Maltose at 0.4 M in the support medium was the most important factor influencing GUS activity in bombarded tissues. GUS activity, 1 day post-bombardment, reached 52 ± 17 GUS-positive foci/MDE (mean ± s.e.m, n=3), with 17 ± 4 foci/MDE at 15 days, giving a 3.0-fold increase (p〈0.05) compared to expression in MDEs bombarded on medium without a high osmoticum treatment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006384525513
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  • 41
    Electronic Resource
    Electronic Resource
    Sorption Kinetics and Transformation of DDT in Sediment (1999)
    Springer
    Water, air & soil pollution 110 (1999), S. 57-66 
    Details
    ISSN: 1573-2932
    Keywords: DDT ; kinetic ; organic pollutant ; sediment ; sorption ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The overall objective of this study was to investigate the sorption kinetics of DDT in sediment under similar experimental conditions employed in corresponding toxicity studies for bentic organisms. A batch of aerated Schoonrewoerdse Wiel sediment, initially spiked with DDT, was sampled over a period of seven days. Concentrations of DDT, DDD and DDE were determined in both the solid and the solution phase in the sediment/water system after separation by centrifugation. It was found that the extractable amount of DDT decreased with increasing contact time. This can partly be explained in terms of transformation of DDT into DDD. Furthermore, the present applied extraction procedure seems to be less effective with increasing contact time, indicating an increase in binding strength of DDT with the sediment material. Finally, on the basis of DDT, DDE and DDD concentrations in both the solid phase and the solution phase, partition coefficients were calculated, which appeared to be independent of the contact time. This points at a very rapid equilibrating between DDT in pore water and in the extractable forms adsorbed at the solid phase.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005087429581
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  • 42
    Electronic Resource
    Electronic Resource
    Environmental Risk Assessment for Pesticides in the Atmosphere; The Results of an International Workshop (1999)
    Springer
    Water, air & soil pollution 115 (1999), S. 5-19 
    Details
    ISSN: 1573-2932
    Keywords: atmospheric fate ; atmospheric transport ; deposition ; emission ; long-range transport ; pesticides ; registration ; remote area ; risk assessment ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract The Health Council of the Netherlands organised an international workshop on the fate of pesticides in the atmosphere and possible approaches for their regulatory environmental risk assessment. Approximately forty experts discussed what is currently known about the atmospheric fate of pesticides and major gaps in our understanding were identified. They favoured a tiered approach for assessing the environmental risks of atmospheric dispersion of these chemicals. In the first tier a pesticide's potential for emission during application, as well as its volatilisation potential should be assessed. Estimates of the former should be based on the application method and the formulation, estimates of the latter on a compound's solubility in water, saturated vapour pressure and octanol/water partition coefficient. Where a pesticide's potential for becoming airborne exceeds critical values, it should be subjected to a more rigorous second tier evaluation which considers its toxicity to organisms in non-target areas. This evaluation can be achieved by calculating and comparing a predicted environmental concentration (PEC) and a predicted no-effect concentration (PNEC). By applying an extra uncertainty factor the PNEC can be provisionally derived from standard toxicity data that is already required for the registration of pesticides. Depending on the distance between the source and the reception area, the PEC can be estimated for remote areas using simple dispersion, trajectory type models and for nearby areas using common dispersion models and standard scenarios of pesticide use. A pesticide's atmospheric transport potential is based on factors such as its reaction rate with OH radicals. It should be used to discriminate between those compounds for which only the risks to nearby ecosystems have to be assessed, and those for which the risks to remote ecosystems also have to be determined. The participants were of the opinion that this approach is, in principle, scientifically feasible, although the remaining uncertainties are substantial. Further field and laboratory research is necessary to gain more reliable estimates of the physico-chemical properties of pesticides, to validate and improve environmental fate models and to validate the applicability of standard toxicity data. This will increase both the accuracy of and our confidence in the outcome of the risk assessment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005238703698
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  • 43
    Electronic Resource
    Electronic Resource
    Institutional transformation at South African universities: Implications for academic staff (1999)
    Springer
    Higher education 38 (1999), S. 275-290 
    Details
    ISSN: 1573-174X
    Keywords: academic staff ; curriculum change ; equity ; governance ; staff development ; student needs ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Nature of Science, Research, Systems of Higher Education, Museum Science
    Notes: Abstract In South Africa the restructuring of the higher education system and the transformation of higher education institutions are located within the country's broad political and socio-economic transition to democracy. This paper focuses particularly on institutional transformation, and pays attention to the implications of the process of transformation for academic staff. The following five interlinked and interdependent issues characterizing institutional transformation in South African higher education are identified: democratising the governance structures of institutions increasing access for educationally and financially disadvantaged students restructuring the curriculum focusing on developmental needs in research and community service redressing inequalities in terms of race and gender. Although the overall effect of institutional transformation is experienced rather negatively by many academic staff members, the paper concludes that academics have to be empowered by means of staff development to remain active partners in the transformation process.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1003768229291
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  • 44
    Electronic Resource
    Electronic Resource
    Airflow and temperature distribution in two-dimensional drying bins (1999)
    Springer
    Journal of engineering mathematics 36 (1999), S. 241-254 
    Details
    ISSN: 1573-2703
    Keywords: flows in porous media ; transformation ; heat transfer ; drying bins ; conformal mapping.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Mathematics , Technology
    Notes: Abstract The design of a drying or cooling store aims to provide an even airflow distribution, when aerated, for preservation purposes. The airflow in some curved bottom bins are studied in this paper. The flow is modelled, using Darcy's law. A generalized Schwarz-Christoffel transformation is employed to reduce the problem of computing streamlines and isobars of airflow to solving a single nonlinear equation for the flow angle along the wall. Corresponding to different bin shapes, a few computed streamlines and isobars of airflow are presented, showing the effect of changing bottom geometries on the air flow. Heat transfer in such bins is also investigated. Based on an analysis of the far field of airflow, finite-height bins are considered. Analytical solutions of the heat conduction equation in terms of streamlines and isobars are obtained.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1004488010532
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  • 45
    Electronic Resource
    Electronic Resource
    Long-Term Transformation and Redistribution of Potentially Toxic Heavy Metals in Arid-Zone Soils: II. Incubation at the Field Capacity Moisture Content (1999)
    Springer
    Water, air & soil pollution 114 (1999), S. 221-250 
    Details
    ISSN: 1573-2932
    Keywords: arid-zone soils ; field capacity ; fractionation ; heavy metals ; kinetics ; redistribution ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract Solid-phase transformation of added Cd, Cu, Cr, Ni, Pb and Zn, in two arid-zone soils incubated in the field capacity moisture regime for one year, were studied. The heavy metals were fractionated into six empirically defined fractions using a selective sequential dissolution (SSD) protocol optimized for arid-zone soils. Each of these fractions was named based on the major soil component targeted for dissolution during the specific SSD step, but it is not assumed that they are mineralogically and chemically totally specific. The transformations of the metals in the two soils incubated at the field capacity regime were compared with those at the moisture saturation regime (Han and Banin, 1997). An initial fast stage of transformation of the soluble metals from the exchangeable (EXC) fraction to the less labile fractions (the carbonate (CARB) fraction for Cd, Pb, Zn, Ni and Cu, and the organic matter (OM) fraction for Cr, and to some extent Cu and Ni) occurred during the fractionation and within one hour after addition. This was followed by a second stage, involving long-term transformation processes of all metals: added Cd was transferred from the EXC into the CARB fraction; added Cr was transferred from the CARB to the OM fraction and Pb was transferred very slowly to the easily reducible oxide (ERO) fraction. Added Cu, Ni and Zn were transferred from the EXC and CARB fractions into the ERO fraction and to some extent OM and RO fractions. In Part I of this series, we reported that during incubation in the saturated moisture regime, Zn and Ni were transferred mainly into the RO and OM fractions. Cadmium, Cr and Pb underwent the same transformation pathways during the slow long-term process, with slightly different rates, in both water regimes. At low levels of addition, the incubated soils moved over one year towards a distribution similar to that of the native soil. At higher levels, the soils still remained removed from the quasi-equilibrium which characterized the native soil, even at the end of one year of incubation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005006801650
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  • 46
    Electronic Resource
    Electronic Resource
    Expansive Visibilization of Work: An Activity-Theoretical Perspective (1999)
    Springer
    Computer supported cooperative work 8 (1999), S. 63-93 
    Details
    ISSN: 1573-7551
    Keywords: activity theory ; action ; transformation ; expansive learning ; intervention ; visibilization ; health care ; medical records
    Source: Springer Online Journal Archives 1860-2000
    Topics: Computer Science
    Notes: Abstract Work is commonly made visible along two dimensions: the linear and the socio-spatial. Both are limited to depicting work in terms of relatively discrete actions. Activity theory introduces the crucial distinction between collective activity systems and individual actions. Expansive visibilization of collective activity systems offers a powerful intervention methodology for dealing with major transformations of work. The linear and the socio-spatial dimensions of work actions are seen in the broader perspective of a third, developmental dimension of work activity. Four steps are identified in a cycle of expansive visibilization, combining activity-level visions and action-level concretizations. The cycle is examined in detail as it unfolded in an intervention study at a children's hospital in Finland. It is concluded that expansive visibilization, driven by contradictions and seeking to reconceptualize the object and motive of work, is not a straightforward process which can be neatly controlled from above. Coherent analytical explanation and goal-setting may come only after the creation and practical implementation of innovative solutions.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008648532192
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  • 47
    Electronic Resource
    Electronic Resource
    Partial male sterility in transgenic tobacco carrying an antisense gene for alternative oxidase under the control of a tapetum-specific promoter (1999)
    Springer
    Molecular breeding 5 (1999), S. 209-218 
    Details
    ISSN: 1572-9788
    Keywords: alternative oxidase ; antisense ; male-sterility ; tapetum-specific promoter ; tobacco ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The alternative oxidase of plant mitochondria is the terminal oxidase of the cyanide-insensitive respiratory pathway and is encoded by a nuclear gene. A 1 kb genomic fragment including exon 3 of the alternative oxidase was amplified by PCR from the genome of Arabidopsis thaliana. This fragment was connected to a tapetum-specific promoter in the antisense orientation and then introduced into tobacco. The pollen viability in three transgenic plants ranged from 2% to 60%. The reduced pollen viability cosegregated with the transgene in a selfed progeny. Immunolocalization of alternative oxidase protein in the immature flower bud section indicated that expression of alternative oxidase protein in tapetum of the transgenic plant was much lower than that of the non-transformant. The histological observation and protein gel-blot analysis showed that the development of pollen grains in the transgenic plant did not progress after the degradation of the tapetum, and the amount of alternative oxidase in pollen grains of the transgenic plant became lower than that of the non-transformant. These results suggested that the alternative oxidase activity in the tapetum has a significant effect on the pollen development.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009606601521
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  • 48
    Electronic Resource
    Electronic Resource
    Transformation of the endospermous legume guar (Cyamopsis tetragonoloba L.) and analysis of transgene transmission (1999)
    Springer
    Molecular breeding 5 (1999), S. 521-529 
    Details
    ISSN: 1572-9788
    Keywords: Cyamopsis tetragonoloba ; β-lactamase inhibitor ; sulbactam ; transformation ; transgene stability ; transgenic guar
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A procedure for transformation of the large-seeded endospermous legume guar (Cyamopsis tetragonoloba L.) and a study on transmission of the transgenes to offspring generations are presented. Using Agrobacterium tumefaciens with a T-DNA construct harbouring a β-glucuronidase gene (uidA) and a neomycin phosphotransferase gene (nptII), maximum transformation frequencies of cotyledonary explants were obtained using 145 mg/l kanamycin sulfate as selective agent. Carbenicillin and cefotaxime, used for the elimination of Agrobacterium after co-culture, displayed considerable toxicity to guar tissues but replacing most of these β-lactams by the non-phytotoxic β-lactamase inhibitor sulbactam as well as addition of thidiazuron and silver thiosulfate increased transformation frequencies up to 10-fold in total. The presence of the transgenes in the primary transformants was demonstrated by genomic DNA analysis of GUS-positive shoots. Chimaeric plants (5–10%) were identified by GUS analysis at the flowering stage and were discarded. Analysis of the R1 offspring from 17 independent transformants showed that in 41% of those, the uidA gene(s) was expressed and stably inherited consistent with Mendelian genetics. This was also found for the R2 and R3 generations of single copy transformants. On the other hand, a large proportion (47%) of the primary transformants gave R1 offspring in which 100% of the plants were GUS-negative. Analysis of these plants by PCR revealed that, at least, most of the transgene sequences were absent, suggesting that they had not been transmitted from the parent transformants. This occurred at similar high frequencies (40–50%) irrespective of the estimated copy number of the transgenes. Thus, major parts of the transgenes, even when present in multiple copies, displayed aberrant transmission, at a high frequency, in the process of going from the primary transformants to the first offspring generation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009602722423
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  • 49
    Electronic Resource
    Electronic Resource
    Expression of the Bar Gene Confers Herbicide Resistance in Transgenic Lettuce (1999)
    Springer
    Transgenic research 8 (1999), S. 33-44 
    Details
    ISSN: 1573-9368
    Keywords: Lactuca sativa ; Agrobacterium tumefaciens ; bialaphos ; phosphinothricin acetyltransferase ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Resistance to bialaphos, a broad-spectrum herbicide, was introduced into Lactuca sativa cv. Evola by Agrobacterium tumefaciens-mediated transformation. A. tumefaciens strains 0310 and 1310, both carrying the bialaphos resistance (bar) and neomycin phosphotransferase (nptII) genes, were used for transformation. Primary transformants were selected on kanamycin sulphate-supplemented shoot regeneration medium. Integration of both transgenes was confirmed by non-radioactive Southern hybridisation. The hypervirulent plasmid ToK47 in A. tumefaciens strain 1310 generated multiple insertions of T-DNA in some transgenic plants; the absence of pToK47 (strain 0310) resulted in single gene inserts in all plants tested. Resistance to glufosinate ammonium was observed in axenic seedlings grown on medium supplemented with the herbicide at 5 mg l−1 and in glasshouse-grown plants sprayed with the compound at 300 mg l−1. Stable expression of the bar gene was observed in R2 generation plants. The kanamycin resistance of R1 seedlings was observed by germinating seeds on medium supplemented with 200 mg l−1 kanamycin sulphate. The presence of NPTII protein and PAT enzyme activity were demonstrated by ELISA and PAT enzyme assay respectively. Transgenes segregated in a Mendelian fashion in some plant lines in the R1 generation; herbicide resistance also segregated in the expected ratio in the R2 generation in most transgenic lines. This study confirmed that an agronomically important transgene can be integrated and stably expressed over several generations in lettuce.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008891216134
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  • 50
    Electronic Resource
    Electronic Resource
    Agrobacterium‐mediated transformation of lavandin (Lavandula x intermedia Emeric ex Loiseleur) (1999)
    Springer
    Transgenic research 8 (1999), S. 335-347 
    Details
    ISSN: 1573-9368
    Keywords: Agrobacterium tumefaciens ; ß‐glucuronidase ; lamiaceae ; lavandin ; neomycin phosphotransferase II ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Lavandin (Lavandula x Emeric ex Loiseleur) is an aromatic plant, the essential oil of which is widely used in the perfume, cosmetic, flavouring and pharmaceutical industries. The qualitative or quantitative modification of its terpenes‐containing essential oil by genetic engineering could have important scientific and commercial applications. In this study, we report the first Agrobacterium tumefaciens‐mediated gene transfer into lavandin. The transformation protocol was optimized by lengthening precultivation and cocultivation periods and by testing five different bacterial strains. We obtained transformed callus lines at a frequency of 40–70 with strains AGL1/GI, EHA105/GI and C58/GI. Transgenic shoots were regenerated from these kanamycin resistant calli and rooted on selective medium with 150 mg l-1 kanamycin. The final percentage of transgenic plants obtained varied from 3 to 9, according to the strain used, within 6 months of culture. The presence of the introduced β‐glucuronidase and neomycin phosphotransferase II genes was shown both by PCR and Southern blot analysis. Transgene expression was investigated using histoenzymatic β‐glucuronidase assays, leaf callus assays and RT‐PCR. Results showed that both β‐glucuronidase and neomycin phosphotransferase II genes were expressed at a high level in at least 41 of the transgenic plants regenerated. This efficient transformation strategy could be used to modify some genetic traits of lavandin (flower colour, pathogens resistance) and to study the biosynthesis of the major monoterpene components of its essential oil (linalool, linalyl acetate, camphor and 1,8‐cineole).
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008948113305
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  • 51
    Electronic Resource
    Electronic Resource
    Simple method for plasmid mediated transformation of different Bacillus species (1999)
    Springer
    Biotechnology techniques 13 (1999), S. 337-340 
    Details
    ISSN: 1573-6784
    Keywords: Bacillus ; plasmids ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract A simple and easy method for the introduction of plasmid DNA into different species of Bacillus was developed. The method involves the suspension in a transformation buffer of nutrient agar grown cells in their late exponential phase and the addition of unpurified plasmid DNA. Transformants were obtained at a frequency of about 103 to 105 stable transformants per μg of plasmid DNA.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1008941007983
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  • 52
    Electronic Resource
    Electronic Resource
    A promoter from sugarcane bacilliform badnavirus drives transgene expression in banana and other monocot and dicot plants (1999)
    Springer
    Plant molecular biology 39 (1999), S. 1221-1230 
    Details
    ISSN: 1573-5028
    Keywords: constitutive expression ; GFP ; GUS ; Musa ; ScBV ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A 1369 bp DNA fragment (Sc) was isolated from a full-length clone of sugarcane bacilliform badnavirus (ScBV) and was shown to have promoter activity in transient expression assays using monocot (banana, maize, millet and sorghum) and dicot plant species (tobacco, sunflower, canola and Nicotiana benthamiana). This promoter was also tested for stable expression in transgenic banana and tobacco plants. These experiments showed that this promoter could drive high-level expression of the β-glucuronidase (GUS) reporter gene in most plant cells. The expression level was comparable to the maize ubiquitin promoter in standardised transient assays in maize. In transgenic banana plants the expression levels were variable for different transgenic lines but was generally comparable with the activities of both the maize ubiquitin promoter and the enhanced cauliflower mosaic virus (CaMV) 35S promoter. The Sc promoter appears to express in a near-constitutive manner in transgenic banana and tobacco plants. The promoter from sugarcane bacilliform virus represents a useful tool for the high-level expression of foreign genes in both monocot and dicot transgenic plants that could be used similarly to the CaMV 35S or maize polyubiquitin promoter.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006125229477
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  • 53
    Electronic Resource
    Electronic Resource
    Changes in morphology, cytoskeleton, and substrate dependence of proliferation after transfection of immortalized rat embryonic fibroblasts with E7 gene of type 16 human papilloma virus (1999)
    Springer
    Bulletin of experimental biology and medicine 127 (1999), S. 99-102 
    Details
    ISSN: 1573-8221
    Keywords: human papilloma virus ; transformation ; actin ; fibronectin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Transfection of rat embryonic fibroblasts with E7 gene of type 16 human papilloma virus changed the cytoskeleton and cell-cell and cell-matrix interactions in two clones of transformed cells. Cell morphology and substrate-dependent proliferation were also changed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02432813
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  • 54
    Electronic Resource
    Electronic Resource
    Transformation of pasta wheat (Triticum turgidum L. var. durum) with high-molecular-weight glutenin subunit genes and modification of dough functionality (1999)
    Springer
    Molecular breeding 5 (1999), S. 377-386 
    Details
    ISSN: 1572-9788
    Keywords: Triticum turgidum L. var. durum ; pasta wheat ; transformation ; seed protein modification ; flour quality improvement
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Particle bombardment has been used to transform three cultivars (L35, Ofanto, Svevo) and one breeding line (Latino × Lira) of durum wheat (Triticum turgidum L. var. durum). These varieties were co-transformed with plasmids containing selectable and scorable marker genes (bar and uidA) and plasmids containing one of two high-molecular-weight (HMW) glutenin subunit genes (encoding subunits 1Ax1 or 1Dx5). Ten independent transgenic lines were recovered from 1683 bombarded scutella (transformation efficiency thus 0.6%). Five lines expressed either subunit 1Dx5 or 1Ax1 at levels similar to those of endogenous subunits encoded on chromosome 1B. To identify the effects of the transgenes on the functional properties of grain, three lines showing segregation for transgene expression were used to isolate sibling T2 plants which were null or positive for the transgene product. Analysis of these plants using a small-scale mixograph showed that expression of the additional subunits resulted in increased dough strength and stability, demonstrating that transformation can be used to modify the quality of durum wheat for bread and pasta making.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009681321708
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    pBIN20: An Improved Binary Vector for shape Agrobacterium-mediated Transformation (1998)
    Springer
    Plant molecular biology reporter 16 (1998), S. 129-131 
    Details
    ISSN: 1572-9818
    Keywords: Agrobacterium tumefaciens ; binary vector ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We report the construction of a binary vector for Agrobacterium tumefaciens-mediated transformation, pBIN20, which contains a superlinker region located between the left and right Ti border sequences. This vector, derived from pBI121, simplifies the cloning of plant expression cassettes and has been used in our laboratory to create lines of transgenic BY-2 tobacco cells. This new vector contains more than 20 unique restriction sites as well as the nptII selectable marker gene within the Ti-DNA borders.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1007444100898
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    Double transformation at the eastern border of the EU: the case of the Euroregion pro Europa Viadrina (1998)
    Springer
    GeoJournal 44 (1998), S. 215-224 
    Details
    ISSN: 1572-9893
    Keywords: cross-border region ; transformation ; regional economic development ; Poland ; Germany
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract One of the major means to foster European integration is the establishment of border spanning regions (‘Euroregions’). This is particularly important on the Eastern borders of the EU, e.g. in Eastern Germany. There, however, a double transformation to post-socialist society is taking place, both inside and outside the EU. Tensions arise between objectives on local and higher political levels, intensified by totally different economic structures and access to EU funds on both sides of the border. This is particularly true for the case of the emerging Euroregion Viadrina. Problems in preserving old industrialised localities in East Germany (e.g. steel) and attempts to resurrect the urban fair place Frankfurt/Oder, clash with transition in agriculture and consumer industries and with new concepts in tourism development and environmental protection in the Polish border zone. In region building, political, economic and ideological goals compete with each other. Local initiatives and higher political governance may both support and hamper each other. The same holds true for the interdependence of cultural integration and economic development. The paper concludes that regional economic development can only be expected if, via the building of the Euroregion, the interplay of these factors leads to compromise and harmonization between the different parties involved.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006818426821
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    The non-agricultural economy in post-socialist rural Romania: The insights and perceptions of national, regional and local institutions (1998)
    Springer
    GeoJournal 46 (1998), S. 199-205 
    Details
    ISSN: 1572-9893
    Keywords: chamber of commerce ; economy ; experts ; institutions ; local government ; Romania ; rural ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract This study evaluates some aspects of the socio-economic transformation of rural Romania with reference to the views of representative organisations (at national, regional and local levels) and other experts. Interviews conducted in ten communes of nine Romanian counties (‘judete’) focus attention on the advantages and disadvantages of system change experienced since 1989; the most important problems and constraints for future socio-economic change; and appropriate policies and perspectives for development in the immediate future. Wherever appropriate the claims of interviewees are substantiated through reference to statistics, drawn in many cases from Chambers of Commerce & Industry (CCI). Local level representatives presented much more negative views on recent change than their national and regional level counterparts, but all agreed on the crucial problem of capital shortage. Thus while specific programmes to assist rural areas are justified, they cannot fully succeed until the national economy is able to grow more rapidly and attract greater foreign investment.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006947932112
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    Transformation of a border region: dispersed agricultural communities in Brasov County, Romania (1998)
    Springer
    GeoJournal 46 (1998), S. 305-317 
    Details
    ISSN: 1572-9893
    Keywords: agriculture ; border region ; communities ; commuting ; conservation ; Romania ; tourism ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract A most distinctive feature of the settlement pattern of the Brasov area is the extreme dispersal of mixed farming encountered in the western extreme of the county to the north and south of Zarnesti: the Bran and Poiana Marului areas. Here a system of peasant subsistence farming developed in a political borderland between the Habsburg and Ottoman Empires. Despite feudal pressures, the peasantry took all available opportunities to extend their independence including elaborate transhumance systems. And after seeing transfrontier commerce as a source of plunder, in the tradition of Balkan highway robbery within relatively unregulated spaces, the peasantry has profited through employment in factories, particularly during the communist period. However, the current recession in manufacturing is throwing the rural population back on limited land resources. Although farming assumes an important subsistence role which contributes to stability, the long-term survival of these communities will depend on new sources of income. Rural tourism has considerable potential and a promising start has been made in Bran. There are, however, constraints on the further development of the business and great attention will have to be given to the conservation of the environment in both the Bucegi Mountains and the Piatra Craiului where national park status is proposed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006984906613
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    Rural diversification and socio-economic transformation in Croatia (1998)
    Springer
    GeoJournal 46 (1998), S. 263-269 
    Details
    ISSN: 1572-9893
    Keywords: Croatia ; decentralisation ; diversification ; industry ; innovation ; rural ; transformation ; urbanisation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Geography
    Notes: Abstract Rural diversification in Croatia is well advanced because many rural families have been able to find work in secondary and tertiary activities without the need to migrate to the towns. Many rural settlements have now attained an urban character although there are regional variations, including a contrast between the continental zone with a relatively high level of commitment to agriculture and the coastal areas, with pronounced ‘deagrarisation’ where the ports and tourist resorts are well developed and the natural resource conditions for agriculture are poor. These variations are examined at the municipality level with reference to two key indicators: the share of nonagricultural population and the share of workers in the total active population. Four categories of socio-economic transformation are recognised: more urbanised, urbanised, less urbanised and rural. The main regional differences between the continental and coastal areas are confirmed with the latter showing a relatively high level of socio-economic transformation through the prominence of more highly urbanised municipalities.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006976704796
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    The insulin-like growth factors and breast cancer — revisited (1998)
    Springer
    Breast cancer research and treatment 47 (1998), S. 197-199 
    Details
    ISSN: 1573-7217
    Keywords: breast cancer ; insulin-like growth factor system ; IGF-I receptor ; IGF-II receptor ; binding proteins ; prognosis ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract In 1992, a special issue of Breast Cancer Research and Treatment was devoted to the insulin-like growth factors and breast cancer. In that issue, identification of the key components of the IGF system was reviewed and their potential role in breast cancer growth was described. In this issue, we revisit the IGF system with particular attention to data that further supports their role in the growth regulation of breast cancer. Several new facets of the IGF system are described, and several laboratories have more clearly defined how each individual component of the IGF system may influence breast cancer biology.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1005938615798
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    Genetic transformation of astaxanthin mutants of Phaffia rhodozyma (1998)
    Springer
    Antonie van Leeuwenhoek 73 (1998), S. 147-153 
    Details
    ISSN: 1572-9699
    Keywords: transformation ; plasmid integration ; Phaffia rhodozyma
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Stable red astaxanthin-producing transformants were obtained after genetic transformation of two Phaffia rhodozyma mutants. A yellow mutant, accumulating β-carotene, and an albino mutant, accumulating phytoene, from P. rhodozyma were transformed using a genomic library of wild-type strain UCD 67-385 in the pBluescript vector. Hybridization assays, using the pBluescript DNA as a radioactive probe, indicate integration of vector sequences into the genome of the transformants. Transformants DNA was digested with restriction endonucleases, ligated with T4 DNA ligase and then used to transform E. coli. Ampicillin resistant plasmids, containing 0.1, 0.2, and 2.5 kb DNA inserts of P. rhodozyma, were rescued from the yeast red transformants. The molecular analysis indicate that transformation has occurred by an integration event of donor DNA into the genome of the host strains.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1000602110765
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    Molecular wheat breeding by direct gene transfer (1998)
    Springer
    Euphytica 100 (1998), S. 219-223 
    Details
    ISSN: 1573-5060
    Keywords: cereals ; wheat ; transformation ; genetic modification ; transgenic plants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A method for efficient genetic transformation of wheat has been developed using immature embryos as targets for microprojectile-mediated gene transfer and a helium driven particle delivery system. Screening and selection of transgenic cells, somatic embryos and regenerated plants are performed with the gus-gene and the phosphinothricin acetyl transferase (bar) gene coding for Basta-resistance as the selectable marker. On average, one fertile transgenic plant can be obtained from about 100 microprojectile treated, immature embryos. The number of integrated copies of the transferred gene ranges from 1 up to about 10. Stable integrated genes are inherited in most of the transgenic lines in a normal mendelian fashion segregating 3:1 in the F2. Homozygous, as well as heterozygous, lines have been followed and analysed genetically at the molecular level and up to F5. Apart from normal stable gene expression, examples have also been found which showed a loss of gene activity or unexpected segregation pattern. For applied aspects, different genes are transferred aiming for improved disease resistance, modification of quality, or other characteristics. First results from these transgenic lines are reported, and problems still existing with the production of stable transgenic wheat lines are discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1018356223333
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    Induction of a virulence response in Agrobacterium tumefaciens by exudates of Pinus pinea cotyledons (1998)
    Springer
    Plant cell, tissue and organ culture 55 (1998), S. 175-181 
    Details
    ISSN: 1573-5044
    Keywords: Conifer ; transformation ; virulence genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract As a preliminary step in efforts to develop a successful protocol for Agrobacterium-mediated transformation of cotyledonary explants of Pinus pinea L. embryos, we tested the ability of embrionary exudates of this species to induce the expression of the virulence genes virA, virB, virC, virD, virE and virG in Agrobacterium tumefaciens containing vir: lacZ fusion constructs. The results obtained in the vir induction assay indicated the absence of bactericidal or bacteriostatic plant compounds affecting A. tumefaciens growth, and showed that cotyledonary and embrionary exudates of P. pinea are able to induce all virulence genes studied, except virG. The data suggest that A. tumefaciens can be used for gene transfer into this important forest and fruit species.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1006295622061
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    Space-time estimation of grid-cell hourly ozone levels for assessment of a deterministic model (1998)
    Springer
    Environmental and ecological statistics 5 (1998), S. 197-222 
    Details
    ISSN: 1573-3009
    Keywords: kriging ; non-separable space-time correlation ; spatial scale ; transformation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Abstract We present an approach to estimate hourly grid-cell surface ozone concentrations based on observations from point monitoring sites in space, for comparison with grid-based results from the SARMAP photochemical air-quality model for a region of northern California. Statistical estimation is carried out on a transformed (square root) scale, followed by back-transforming to the original scale of ozone in parts per billion, adjusting for bias and variance. We estimate a spatially-varying diurnal mean structure and a non-separable space-time correlation structure on the transformed scale. Temporal pre-whitening is followed by modelling of a spatially non-stationary, diurnally-varying spatial correlation structure using a spatial deformation approach. Comparisons of SARMAP model results with the estimated grid-cell ozone levels are presented.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009663518685
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    Characterization of receptors with a new negative image: Use in molecular docking and lead optimization (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 321-336 
    Details
    ISSN: 0887-3585
    Keywords: surface characterization ; DOCK ; structure-based molecular design ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The characterization of receptor binding sites is an important aspect of molecular docking, molecular recognition, and the structure-based design process. This characterization can take several forms: the receptor surface itself can be delineated or described, the space adjacent to the surface can be chemically mapped, or a negative image of the protein binding region can be generated. In this report, we describe a new method of constructing a negative image through generation of a set of spheres. These spheres lie along the receptor surface, and their centers represent possible ligand atom positions. By the method in which they are constructed, these spheres carry a limited amount of energetic and chemical information in addition to their primary geometric information. We test the accuracy of the image by comparing sphere positions to the positions of bound ligand atoms and propose a figure of merit for such tests. Then, we use the spheres to orient ligands in enzyme active sites and show how they can be used to generate low scoring configurations more efficiently than other approaches that search orientation space. In addition, two novel applications of these spheres are described: they are used to help identify structural differences among families of enzymes and to suggest points for ligand modification in analog design. Proteins 30:321-336, 1998. © 1998 Wiley-Liss, Inc.
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    Nitric oxide myoglobin: Crystal structure and analysis of ligand geometry (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 352-356 
    Details
    ISSN: 0887-3585
    Keywords: myoglobin ; nitric oxide ; ligand binding ; X-ray crystallography ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The structure of the ferrous nitric oxide form of native sperm whale myoglobin has been determined by X-ray crystallography to 1.7 Å resolution. The nitric oxide ligand is bent with respect to the heme plane: the Fe-N-O angle is 112°. This angle is smaller than those observed in model compounds and in lupin leghemoglobin. The exact angle appears to be influenced by the strength of the proximal bond and hydrogen bonding interactions between the distal histidine and the bound ligand. Specifically, the Nε atom of histidine64 is located 2.8 Å away from the nitrogen atom of the bound ligand, implying electrostatic stabilization of the FeNO complex. This interpretation is supported by mutagenesis studies. When histidine64 is replaced with apolar amino acids, the rate of nitric oxide dissociation from myoglobin increases tenfold. Proteins 30:352-356, 1998. © 1998 Wiley-Liss, Inc.
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    A general method of domain closure is applied to phosphoglycerate kinase and the result compared with the crystal structure of a closed conformation of the enzyme (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 372-380 
    Details
    ISSN: 0887-3585
    Keywords: protein modeling ; crystal structure ; conformation change ; prediction ; mechanism ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The occurrence of large domain motions associated with the mechanism of action of many proteins is well established. We present a general method of predicting domain closure applicable to proteins containing domains separated by an apparent hinge. The method attempts to allow for natural directional bias within the closing protein by repeatedly applying a weak pulling force over a short distance between pairs of atoms chosen at random in the two domains in question. Appropriate parameters governing the pulling function were determined empirically. The method was applied to the bi-lobal protein PGK and a closed-form activated ternary complex generated for Bacillus stearothermophilus PGK. This model was compared with the recently determined crystal structure of closed-form Trypanosoma brucei PGK. The model predicts the correct hinge regions, although the magnitude of movement at one hinge point was overestimated, and provides a reasonable representation of the closed-form ternary complex. Proteins 30:372-380, 1998. © 1998 Wiley-Liss, Inc.
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    Improved protein free energy calculation by more accurate treatment of nonbonded energy: Application to chymotrypsin inhibitor 2, V57A (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 388-400 
    Details
    ISSN: 0887-3585
    Keywords: molecular dynamics ; free energy perturbation ; thermodynamics integration ; spherical solvent boundary potential ; cell multipole method ; Nosé-Hoover equation ; component analysis ; chymotrypsin inhibitor 2 ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: We developed a software package for improved free energy calculation, in which spherical solvent boundary potential, cell multipole method, and Nosé-Hoover equation are employed. The performance of the developed software package is demonstrated in the case of valine to alanine mutation of the 57th residue in chymotrypsin inhibitor 2. By using this package, we obtained results quantitatively comparable to experimental results. By the free energy component analysis, it is shown that leucine 51, arginine 65, arginine 67, and phenylalanine 69 residues contribute significantly to the total free energy shift, ΔΔG. Among them, contribution from the hydrophilic arginine 67 residue, which is in close contact with the mutation site, is the largest. Structure around the mutation site is largely changed by the mutation. The structure change is caused mainly by two effects, hydrophobic interaction and short-range interaction along the sequence. Effects of Nosé-Hoover algorithm and Kirkwood reaction field are also discussed. Proteins 30:388-400, 1998. © 1998 Wiley-Liss, Inc.
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    Proteolysis as a probe of thermal unfolding of cytochrome C (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 435-441 
    Details
    ISSN: 0887-3585
    Keywords: cytochrome c ; thermal unfolding ; proteolysis ; proteinase K ; thermolysin ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Recent hydrogen exchange experiments on native cytochrome c implicate a sequential unfolding pathway in contrast to a simple two-state process. We have studied the heat-induced unfolding of this protein by using spectroscopic measurements to detect changes in conformation and proteolytic enzyme digestion to identify regions of the protein that are labile. Several spectroscopic profiles were monitored: CD at 222 nm, a measurement of secondary structure change in the protein, the absorbance at 280 nm, involving the local environment of Trp 59, and absorbance at 420 nm, the Soret band of the heme. The apparent Tm values for these probes differ, consistent with an unfolding pathway containing intermediates. The limited digestion by proteinase K is consistent with population of an intermediate state in unfolding. We find a single strong region of cleavage at low temperature with retention of structure in each fragment. Proteins 30:435-441, 1998. © 1998 Wiley-Liss, Inc.
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    Crystallographic and spectroscopic characterization of a molecular hinge: Conformational changes in bothropstoxin I, a dimeric Lys49-phospholipase A2 homologue (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 442-454 
    Details
    ISSN: 0887-3585
    Keywords: venom toxin ; protein-membrane interaction ; X-ray diffraction ; spectroscopy ; quaternary structural change ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Bothropstoxin I (BthTX-I) from the venom of Bothrops jararacussuis a myotoxic phospholipase A2 (PLA2) homologue which, although catalytically inactive due to an Asp49→Lys substitution, disrupts the integrity of lipid membranes by a Ca2+-independent mechanism. The crystal structures of two dimeric forms of BthTX-I which diffract X-rays to resolutions of 3.1 and 2.1 Å have been determined. The monomers in both structures are related by an almost perfect twofold axis of rotation and the dimer interfaces are defined by contacts between the N-terminal α-helical regions and the tips of the β-wings of partner monomers. Significant differences in the relative orientation of the monomers in the two crystal forms results in “open” and “closed” dimer conformations. Spectroscopic investigations of BthTX-I in solution have correlated these conformational differences with changes in the intrinsic fluorescence emission of the single tryptophan residues located at the dimer interface. The possible relevance of this structural transition in the Ca2+-independent membrane damaging activity is discussed. Proteins 30:442-454, 1998. © 1998 Wiley-Liss, Inc.
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    Structure-based design of model proteins (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 10-20 
    Details
    ISSN: 0887-3585
    Keywords: protein design ; lattice model ; protein stability ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A structure-based, sequence-design procedure is proposed in which one considers a set of decoy structures that compete significantly with the target structure in being low energy conformations. The decoy structures are chosen to have strong overlaps in contacts with the putative native state. The procedure allows the design of sequences with large and small stability gaps in a random-bond heteropolymer model in both two and three dimensions by an appropriate assignment of the contact energies to both the native and nonnative contacts. The design procedure is also successfully applied to the two-dimensional HP model. Proteins 31:10-20, 1998. © 1998 Wiley-Liss, Inc.
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    Modelling repressor proteins docking to DNA (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 535-549 
    Details
    ISSN: 0887-3585
    Keywords: docking ; protein-DNA ; prediction ; structure ; base recognition ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The docking of repressor proteins to DNA starting from the unbound protein and model-built DNA coordinates is modeled computationally. The approach was evaluated on eight repressor/DNA complexes that employed different modes for protein/ DNA recognition. The global search is based on a protein-protein docking algorithm that evaluates shape and electrostatic complementarity, which was modified to consider the importance of electrostatic features in DNA-protein recognition. Complexes were then ranked by an empirical score for the observed amino acid /nucleotide pairings (i.e., protein-DNA pair potentials) derived from a database of 20 protein/DNA complexes. A good prediction had at least 65% of the correct contacts modeled. This approach was able to identify a good solution at rank four or better for three out of the eight complexes. Predicted complexes were filtered by a distance constraint based on experimental data defining the DNA footprint. This improved coverage to four out of eight complexes having a good model at rank four or better. The additional use of amino acid mutagenesis and phylogenetic data defining residues on the repressor resulted in between 2 and 27 models that would have to be examined to find a good solution for seven of the eight test systems. This study shows that starting with unbound coordinates one can predict three-dimensional models for protein/DNA complexes that do not involve gross conformational changes on association. Proteins 33:535-549, 1998. © 1998 Wiley-Liss, Inc.
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    Influence of protein structure databases on the predictive power of statistical pair potentials (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 139-149 
    Details
    ISSN: 0887-3585
    Keywords: protein structure ; statistical potentials ; protein structure database ; assessing protein models ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A long standing goal in protein structure studies is the development of reliable energy functions that can be used both to verify protein models derived from experimental constraints as well as for theoretical protein folding and inverse folding computer experiments. In that respect, knowledge-based statistical pair potentials have attracted considerable interests recently mainly because they include the essential features of protein structures as well as solvent effects at a low computing cost. However, the basis on which statistical potentials are derived have been questioned. In this paper, we investigate statistical pair potentials derived from protein three-dimensional structures, addressing in particular questions related to the form of these potentials, as well as to the content of the database from which they are derived. We have shown that statistical pair potentials depend on the size of the proteins included in the database, and that this dependence can be reduced by considering only pairs of residue close in space (i.e., with a cutoff of 8 Å). We have shown also that statistical potentials carry a memory of the quality of the database in terms of the amount and diversity of secondary structure it contains. We find, for example, that potentials derived from a database containing α-proteins will only perform best on α-proteins in fold recognition computer experiments. We believe that this is an overall weakness of these potentials, which must be kept in mind when constructing a database. Proteins 31:139-149, 1998. © 1998 Wiley-Liss, Inc.
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    Crystal structure of the disulfide-stabilized Fv fragment of anticancer antibody B1: Conformational influence of an engineered disulfide bond (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 128-138 
    Details
    ISSN: 0887-3585
    Keywords: antibody ; antitumor ; single chain Fv ; variable domains ; X-ray crystallography ; protein structure ; protein stability ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A recombinant Fv construct of the B1 monoclonal antibody that recognizes the LewisY-related carbohydrate epitope on human carcinoma cells has been prepared. The Fv is composed of the polypeptide chains of the VH and VL domains expressed independently and isolated as inclusion bodies. The Fv is prepared by combining and refolding equimolar amounts of guanidine chloride solubilized inclusion bodies. The Fv is stabilized by an engineered interchain disulfide bridge between residues VL100 and VH44. This construct has a similar binding affinity as that of the single-chain construct (Benhar and Pastan, Clin. Cancer Res. 1:1023-1029, 1995). The B1 disulfide-stabilized Fv (B1dsFv) crystallizes in space group P6122 with the unit cell parameters a = b = 80.1 Å, and c = 138.1 Å. The crystal structure of the B1dsFv has been determined at 2.1-Å resolution using the molecular replacement technique. The final structure has a crystallographic R-value of 0.187 with a root mean square deviation in bond distance of 0.014 Å and in bond angle of 2.74°. Comparisons of the B1dsFv structure with known structures of Fv regions of other immunoglobulin fragments shows closely related secondary and tertiary structures. The antigen combining site of B1dsFv is a deep depression 10-Å wide and 17-Å long with the walls of the depression composed of residues, many of which are tyrosines, from complementarity determining regions L1, L3, H1, H2, and H3. Model building studies indicate that the LewisY tetrasaccharide, Fuc-Gal-Nag-Fuc, can be accommodated in the antigen combining site in a manner consistent with the epitope predicted in earlier biochemical studies (Pastan, Lovelace, Gallo, Rutherford, Magnani, and Willingham, Cancer Res. 51:3781-3787, 1991). Thus, the engineered disulfide bridge appears to cause little, if any, distortion in the Fv structure, making it an effective substitute for the B1 Fab. Proteins 31:128-138, 1998. Published 1998 Wiley-Liss, Inc.This article is a US Government work and, as such, is in the public domain in the United States of America.
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    Interaction of transmembrane helices by a knobs-into-holes packing characteristic of soluble coiled coils (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 150-159 
    Details
    ISSN: 0887-3585
    Keywords: photosynthetic reaction center ; bacteriorhodopsin ; cytochrome C oxidase ; zipper ; packing ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Membrane-embedded protein domains frequently exist as α-helical bundles, as exemplified by photosynthetic reaction centers, bacteriorhodopsin, and cytochrome C oxidase. The sidechain packing between their transmembrane helices was investigated by a nearest-neighbor analysis which identified sets of interfacial residues for each analyzed helix-helix interface. For the left-handed helix-helix pairs, the interfacial residues almost exclusively occupy positions a, d, e, or g within a heptad motif (abcdefg) which is repeated two to three times for each interacting helical surface. The connectivity between the interfacial residues of adjacent helices conforms to the knobs-into-holes type of sidechain packing known from soluble coiled coils. These results demonstrate on a quantitative basis that the geometry of sidechain packing is similar for left-handed helix-helix pairs embedded in membranes and coiled coils of soluble proteins. The transmembrane helix-helix interfaces studied are somewhat less compact and regular as compared to soluble coiled coils and tolerate all hydrophobic amino acid types to similar degrees. The results are discussed with respect to previous experimental findings which demonstrate that specific interactions between transmembrane helices are important for membrane protein folding and/or oligomerization. Proteins 31:150-159, 1998. © 1998 Wiley-Liss, Inc.
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    Protein folding in the landscape perspective: Chevron plots and non-arrhenius kinetics (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 2-33 
    Details
    ISSN: 0887-3585
    Keywords: chevron plot ; energy landscape ; folding funnel ; kinetic trap ; lattice models ; non-Arrhenius behavior ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: We use two simple models and the energy landscape perspective to study protein folding kinetics. A major challenge has been to use the landscape perspective to interpret experimental data, which requires ensemble averaging over the microscopic trajectories usually observed in such models. Here, because of the simplicity of the model, this can be achieved. The kinetics of protein folding falls into two classes: multiple-exponential and two-state (single-exponential) kinetics. Experiments show that two-state relaxation times have “chevron plot” dependences on denaturant and non-Arrhenius dependences on temperature. We find that HP and HP+ models can account for these behaviors. The HP model often gives bumpy landscapes with many kinetic traps and multiple-exponental behavior, whereas the HP+ model gives more smooth funnels and two-state behavior. Multiple-exponential kinetics often involves fast collapse into kinetic traps and slower barrier climbing out of the traps. Two-state kinetics often involves entropic barriers where conformational searching limits the folding speed. Transition states and activation barriers need not define a single conformation; they can involve a broad ensemble of the conformations searched on the way to the native state. We find that unfolding is not always a direct reversal of the folding process. Proteins 30:2-33, 1998. © 1998 Wiley-Liss, Inc.
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    Feature-extraction from endopeptidase cleavage sites in mitochondrial targeting peptides (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 49-60 
    Details
    ISSN: 0887-3585
    Keywords: kohonen network ; mitochondrial processing peptidase (MPP) ; mitochondrial intermediate peptidase (MIP) ; neural network ; protein import ; sequence motif ; mitochondrial targeting ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Cleavage sites in nuclear-encoded mitochondrial protein targeting peptides (mTPs) from mammals, yeast, and plants have been analysed for characteristic physicochemical features using statistical methods, perceptrons, multilayer neural networks, and self-organizing feature maps. Three different sequence motifs were found, revealing loosely defined arginine motifs with Arg in positions -10, -3, and -2. A self-organizing feature map was able to cluster these three types of endopeptidase target sites but did not identify any species-specific characteristics in mTPs. Neural networks were used to define local sequence features around precursor cleavage sites. Proteins 30:49-60, 1998. © 1998 Wiley-Liss, Inc.
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    Don Caspar, TMV, and protein-protein interactions (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 109-112 
    Details
    ISSN: 0887-3585
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: No abstract.
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    A hybrid sequence approach to the paracelsus challenge (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 136-143 
    Details
    ISSN: 0887-3585
    Keywords: protein design ; protein structure ; circular dichroism ; trifluoroethanol ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Inspired by the Paracelsus Challenge of Rose and Creamer (Proteins 19:1-3, 1994), we have designed a protein sequence that is 50% identical to an all-helical protein but is intended to fold into a largely β-sheet structure. Rather than attempt a de novo design, our strategy was to construct a hybrid sequence based on a helical “parent” protein (434 Cro) and a “target” protein with the desired fold (the B1 domain of protein G). The hybrid sequence (Crotein-G) is 50% identical to 434 Cro but is also 62% identical to the B1 domain of protein G. We also created a variant of Crotein-G (ZCrotein-G) that contains a potential His3Cys1 zinc binding site. At low protein concentrations and in the presence of 20% 2,2,2-trifluoroethanol (TFE) (v/v), the circular dichroism spectra of the designed proteins are distinct from that of 434 Cro and similar to that of the B1 domain of protein G. However, the proteins fail to denature in a cooperative manner. Furthermore, aggregation occurs at moderate protein concentrations or in the absence of TFE. Addition of zinc to ZCrotein-G does not promote structure formation. In summary, 434 Cro has been altered to something that may resemble the B1 domain of protein G, but the protein does not adopt a native structure. Proteins 30:136-143, 1998. © 1998 Wiley-Liss, Inc.
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    Monte Carlo simulation of denaturation of adsorbed proteins (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 168-176 
    Details
    ISSN: 0887-3585
    Keywords: denaturation kinetics ; irreversible conformational changes ; metastable states ; folding temperature ; lattice model ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Denaturation of model proteinlike molecules at the liquid-solid interface is simulated over a wide temperature range by employing the lattice Monte Carlo technique. Initially, the molecule containing 27 monomers of two types (A and B) is assumed to be adsorbed in the native folded state (a 3 × 3 × 3 cube) so that one of its sides is in contact with the surface. The details of the denaturation kinetics are found to be slightly dependent on the choice of the side, but the main qualitative conclusions hold for all the sides. In particular, the kinetics obey approximately the conventional first-order law at T 〉 Tc (Tc is the collapse temperature for solution). With decreasing temperature, below Tc but above Tf (Tf is the folding temperature for solution), deviations appear from the first-order kinetics. For the most interesting temperatures, that is, below Tf, the denaturation kinetics are shown to be qualitatively different from the conventional ones. In particular, the denaturation process occurs via several intermediate steps due to trapping in metastable states. Mathematically, this means that (i) the transition to the denatured state of a given molecule is nonexponential, and (ii) the denaturation process cannot be described by a single rate constant kr. One should rather introduce a distribution of values of this rate constant (different values of kr correspond to the transitions to the altered state via different metastable states). Proteins 30:168-176, 1998. © 1998 Wiley-Liss, Inc.
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    Monte Carlo simulation of the kinetics of protein adsorption (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 177-182 
    Details
    ISSN: 0887-3585
    Keywords: adsorption ; irreversible conformational changes ; denaturation rate constant ; kinetic control ; diffusion control ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Adsorption of proteins occurs via diffusion toward the interface, actual adsorption, and subsequent irreversible conformational changes resulting in denaturation of the native protein structure. The conventional kinetic models describing these steps are based on the assumption that the denaturation transitions obey the first-order law with a single value of the denaturation rate constant kr. Meanwhile, recent Monte Carlo simulations indicate that, in general, the denaturation process cannot be described by a single rate constant kr. One should rather introduce a distribution of this rate constant (physically, different values of kr correspond to the transitions to the altered state via different metastable states). We have calculated the kinetics of irreversible adsorption of proteins with and without distribution of the denaturation rate constant kr in the limits when protein diffusion in the solution is, respectively, rapid or slow. In both cases, the adsorption kinetics with distribution of kr are found to be close to those with a single-valued rate constant kr provided that the average value of kr in the former case is equal to kr for the latter case. This conclusion holds even for wide distributions of kr. The consequences of this finding for the fitting of global experimental kinetics on the basis of phenomenological equations are briefly discussed. Proteins 30:177-182, 1998. © 1998 Wiley-Liss, Inc.
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    Structural diversity of sequentially identical subsequences of proteins: Identical octapeptides can have different conformations (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 228-231 
    Details
    ISSN: 0887-3585
    Keywords: protein folding ; local vs. non-local interactions ; secondary structure prediction ; fragment matching algorithms ; PDB ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: One of the most important questions in the protein folding problem is whether secondary structures are formed entirely by local interactions. One way to answer this question is to compare identical subsequences of proteins to see if they have identical structures. Such an exercise would also reveal a lower limit on the number of amino acids needed to form unique secondary structures. In this context, we have searched the April 1996 release of the Protein Data Bank for sequentially identical subsequences of proteins and compared their structures. We find that identical octamers can have different conformations. In addition, there are several examples of identical heptamers with different conformations, and the number of identical hexamers with different conformations has increased since the previous PDB releases. These observations imply that secondary structure can be formed entirely by non-local interactions and that an identical match of up to eight amino acids may not imply structural similarity. In addition to the larger context of the protein folding problem, these observations have implications for protein structure prediction methods. Proteins 30:228-231, 1998. © 1998 Wiley-Liss, Inc.
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    Structural modeling of the complex between an acetylcholine receptor-mimicking antibody and its snake toxin antigen (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 249-263 
    Details
    ISSN: 0887-3585
    Keywords: antibody-antigen complex ; snake toxin ; protein docking ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The antibody Mα2-3 neutralizes the functional, acetylcholine receptor binding activity of its antigen, neurotoxin α, and exhibits several other properties in common with the receptor itself. We present here the results of calculations examining the three-dimensional structure of the toxin α:Mα2-3 complex. The antigen structure, determined by nuclear magnetic resonance spectroscopy,1 was docked to models of the variable fragment of the antibody combining site2 by using a method based on surface complementarity and maximization of buried surface area3,4 while taking into account the possibility of conformational change on complexation. Extensive experimental information on the location of the functional epitope was incorporated into the analysis and used to screen candidate geometries of the complex resulting from the modeling. Eight plausible structures that are in accord with the experimental data were derived. Common structural features of the models are discussed, and residues of the antibody-combining site that are expected to play important roles in complexation are identified. In particular, three epitope residues that, according to mutagenesis experiments, make particularly strong contributions to the binding, interact excentrically and do not make contact with the central loops of the combining site, L3 and H3. Proteins 30:249-263, 1998. © 1998 Wiley-Liss, Inc.
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    Tertiary structure prediction of the KIX domain of CBP using Monte Carlo simulations driven by restraints derived from multiple sequence alignments (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 287-294 
    Details
    ISSN: 0887-3585
    Keywords: protein structure prediction ; side chain contact prediction ; lattice protein models ; CREB-binding protein ; KIX domain ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Using a recently developed protein folding algorithm, a prediction of the tertiary structure of the KIX domain of the CREB binding protein is described. The method incorporates predicted secondary and tertiary restraints derived from multiple sequence alignments in a reduced protein model whose conformational space is explored by Monte Carlo dynamics. Secondary structure restraints are provided by the PHD secondary structure prediction algorithm that was modified for the presence of predicted U-turns, i.e., regions where the chain reverses global direction. Tertiary restraints are obtained via a two-step process: First, seed side-chain contacts are identified from a correlated mutation analysis, and then, a threading-based algorithm expands the number of these seed contacts. Blind predictions indicate that the KIX domain is a putative three-helix bundle, although the chirality of the bundle could not be uniquely determined. The expected root-mean-square deviation for the correct chirality of the KIX domain is between 5.0 and 6.2 Å. This is to be compared with the estimate of 12.9 Å that would be expected by a random prediction, using the model of F. Cohen and M. Sternberg (J. Mol. Biol. 138:321-333, 1980). Proteins 30:287-294, 1998. © 1998 Wiley-Liss, Inc.
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    Mass spectrometric identification and microcharacterization of proteins from electrophoretic gels: Strategies and applications (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 74-89 
    Details
    ISSN: 0887-3585
    Keywords: mass spectrometry ; matrix-assisted laser desorption/ionization ; electrospray ; database searching ; gel electrophoresis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The entire genomic DNA sequences of a number of prokaryotic and eukaryotic species are now available and many more, including the human genome, will be completed in the near future. The state-of-life of a cell at any given time, however, is defined by its protein composition, i.e., its proteome. Gel electrophoresis, mass spectrometry, and bioinformatics will be important tools for protein and proteome analysis in the post-genome era. Protein identification from electrophoretic gels by mass spectrometric peptide mapping or peptide sequencing combined with sequence database searching is established and has been applied to numerous biological systems. We describe current strategies and selected applications in molecular and cell biology. The next challenges are detailed structure/function analyses, which include studying the molecular composition of multiprotein complexes and characterization of secondary modifications of proteins. The advantages and limitations of a number of mass spectrometry-based strategies designed for microcharacterization of low amounts of protein from electrophoretic gels are discussed and illustrated by examples. Proteins Suppl. 2:74-89, 1998. © 1998 Wiley-Liss, Inc.
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    A fusion protein between serum amyloid A and staphylococcal nuclease - synthesis, purification, and structural studies (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 381-387 
    Details
    ISSN: 0887-3585
    Keywords: serum amyloid A ; fluorescence ; circular dichroism ; acute phase ; denaturation ; nuclease ; amyloidosis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: We developed a recombinant DNA system to overexpress a fusion protein between the small, minimally soluble acute phase serum protein, serum amyloid A (SAA), and the bacterial enzyme staphylococcal nuclease (SN). This fusion protein is very soluble and is immunoreactive to polyclonal anti-SAA antibodies. Tryptophan fluorescence shows smooth denaturation curves for the fusion protein in guanidinium HCl or potassium thiocyanate. Fluorescence also indicates that only a single tryptophan residue (of the four present) is accessible to iodide quenching and, presumably, is exposed on the surface of the fusion protein. Circular dichroism (CD) shows a significant signal indicating α-helix, which can be attributed to the SAA portion of the molecule; these are the first CD spectral data available for SAA. pH titration shows persistence of helix domains for the fusion protein at pH 3.0, in contrast to the denaturation of SN under the same conditions. (The entire fusion protein shows a random coil pattern below pH 3.0.) By exploiting the structural and solubility properties of SN, this fusion protein has provided the first structural data about SAA - the precursor of the amyloid deposits in secondary amyloidosis. This fusion protein should be useful for further physical and physiologic studies of SAA. Proteins 30:381-387, 1998. © 1998 Wiley-Liss, Inc.
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    The PDP: Past, present, and future (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 30 (1998), S. 1-1 
    Details
    ISSN: 0887-3585
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: No abstract.
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    Direct observation of an altered quaternary-structure transition in a mutant aspartate transcarbamoylase (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 383-390 
    Details
    ISSN: 0887-3585
    Keywords: time-resolved small-angle X-ray scattering ; allosterism ; domain closure ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Time-resolved small-angle X-ray scattering (TR-SAXS) was used to monitor the structural changes that occur upon the binding of the natural substrates to a mutant version of the allosteric enzyme aspartate transcarbamoylase from Escherichia coli, in which the creation of a critical link stabilizing the R state of the enzyme is hindered. Previously, SAXS experiments at equilibrium showed that the structures of the unligated mutant enzyme and the mutant enzyme saturated with a bisubstrate analog are indistinguishable from the T and R state structures, respectively, of the wild-type enzyme (Tauc et al., Protein Sci. 3:1998-2004, 1994). However, as opposed to the wild-type enzyme, the combination of one substrate, carbamoyl phosphate, and succinate, an analog of aspartate, did not convert the mutant enzyme into the R state. By using TR-SAXS we have been able to study the transient steady-state during catalysis using the natural substrates rather than the nonreactive substrate analogs. The steady-state in the presence of saturating amount of substrates is a mixture of 60% T and 40% R structures, which is further converted entirely to R in the additional presence of ATP. These results provide a structural explanation for the reduced cooperativity observed with the mutant enzyme as well as for the stimulation by ATP at saturating concentrations of substrates. They also illustrate the crucial role played by domain motions and quaternary-structure changes for both the homotropic and heterotropic aspects of allostery. Proteins 31:383-390, 1998. © 1998 Wiley-Liss, Inc.
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    Structural investigation of C4b-binding protein by molecular modeling: Localization of putative binding sites (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 391-405 
    Details
    ISSN: 0887-3585
    Keywords: complement control protein ; protein modeling ; blood coagulation ; C4b-binding protein ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: C4b-binding protein (C4BP) contributes to the regulation of the classical pathway of the complement system and plays an important role in blood coagulation. The main human C4BP isoform is composed of one β-chain and seven α-chains essentially built from three and eight complement control protein (CCP) modules, respectively, followed by a nonrepeat carboxy-terminal region involved in polymerization of the chains. C4BP is known to interact with heparin, C4b, complement factor I, serum amyloid P component, streptococcal Arp and Sir proteins, and factor VIII/VIIIa via its α-chains and with protein S through its β-chain. The principal aim of the present study was to localize regions of C4BP involved in the interaction with C4b, Arp, and heparin. For this purpose, a computer model of the 8 CCP modules of C4BP α-chain was constructed, taking into account data from previous electron microscopy (EM) studies. This structure was investigated in the context of known and/or new experimental data. Analysis of the α-chain model, together with monoclonal antibody studies and heparin binding experiments, suggests that a patch of positively charged residues, at the interface between the first and second CCP modules, plays an important role in the interaction between C4BP and C4b/Arp/Sir/heparin. Putative binding sites, secondary-structure prediction for the central core, and an overall reevaluation of the size of the C4BP molecule are also presented. An understanding of these intermolecular interactions should contribute to the rational design of potential therapeutic agents aiming at interfering specifically some of these protein-protein interactions. Proteins 31:391-405, 1998. © 1998 Wiley-Liss, Inc.
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    Species dependence of enzyme-substrate encounter rates for triose phosphate isomerases (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 406-416 
    Details
    ISSN: 0887-3585
    Keywords: electrostatics ; Brownian dynamics ; triose phosphate isomerase ; diffusion-control ; similarity index ; rate enhancement ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Triose phosphate isomerase (TIM) is a diffusion-controlled enzyme whose rate is limited by the diffusional encounter of the negatively charged substrate glyceraldehyde 3-phosphate (GAP) with the homodimeric enzyme's active sites. Translational and orientational steering of GAP toward the active sites by the electrostatic field of chicken muscle TIM has been observed in previous Brownian dynamics (BD) simulations. Here we report simulations of the association of GAP with TIMs from four species with net charges at pH 7 varying from -12e to +12e. Computed second-order rate constants are in good agreement with experimental data. The BD simulations and computation of average Boltzmann factors of substrate-protein interaction energies show that the protein electrostatic potential enhances the rates for all the enzymes. There is much less variation in the computed rates than might be expected on the basis of the net charges. Comparison of the electrostatic potentials by means of similarity indices shows that this is due to conservation of the local electrostatic potentials around the active sites which are the primary determinants of electrostatic steering of the substrate. Proteins 31:406-416, 1998. © 1998 Wiley-Liss, Inc.
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    Functional conformational changes of endo-1,4-xylanase II from Trichoderma reesei: A molecular dynamics study (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 434-444 
    Details
    ISSN: 0887-3585
    Keywords: hinge ; structural change ; xylose ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Recent crystallographic studies have revealed a range of structural changes in the three-dimensional structure of endo-1,4-xylanase (XYNII) from Trichoderma reesei. The observed conformational changes can be described as snapshots of an open-close movement of the active site of XYNII. These structures were further analyzed in this study. In addition, a total of four 1 ns molecular dynamics (MD) simulations were performed representing different states of the enzyme. A comparison of the global and local changes found in the X-ray structures and the MD runs suggested that the simulations reproduced a similar kind of active site opening and closing as predicted by the crystal structures. The open-close movement was characterized by the use of distance difference matrixes and the Hingefind program (Wriggers and Schulten, Proteins 29:1-14, 1997) to be a ‘hinge-bending’ motion involving two large rigidly-moving regions and an extended hinge. This conformational feature is probably inherent to this molecular architecture and probably plays a role in the function of XYNII. Proteins 31:434-444, 1998. © 1998 Wiley-Liss, Inc.
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    Interaction of human SRY protein with DNA: A molecular dynamics study (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 417-433 
    Details
    ISSN: 0887-3585
    Keywords: molecular dynamics ; sex-determining region Y (SRY) protein ; high mobility group (HMG) box ; DNA-binding proteins ; DNA bending ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Molecular dynamics simulations have been conducted to study the interaction of human sex-determining region Y (hSRY) protein with DNA. For this purpose, simulations of the hSRY high mobility group (HMG) domain (hSRY-HMG) with and without its DNA target site, a DNA octamer, and the DNA octamer alone have been carried out, employing the NMR solution structure of hSRY-HMG-DNA complex as a starting model. Analyses of the simulation results demonstrated that the interaction between hSRY and DNA was hydrophobic, just a few hydrogen bonds and only one water molecule as hydrogen-bonding bridge were observed at the protein-DNA interface. These two hydrophobic cores in the hSRY-HMG domain were the physical basis of hSRY-HMG-DNA specific interaction. They not only maintained the stability of the complex, but also primarily caused the DNA deformation. The salt bridges formed between the positive-charged residues of hSRY and phosphate groups of DNA made the phosphate electroneutral, which was advantageous for the deformation of DNA and the formation of a stable complex. We predicted the structure of hSRY-HMG domain in the free state and found that both hSRY and DNA changed their conformations to achieve greater complementarity of geometries and properties during the binding process; that is, the protein increased the angle between its long and short arms to accommodate the DNA, and the DNA became bent severely to adapt to the protein, although the conformational change of DNA was more severe than that of the hSRY-HMG domain. The sequence specificity and the role of residue Met9 are also discussed. Proteins 31:417-433, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 12 Ill.
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  • 93
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    Density functional studies on herpes simplex virus type 1 thymidine kinase-substrate interactions: The role of Tyr-172 and Met-128 in thymine fixation (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 453-459 
    Details
    ISSN: 0887-3585
    Keywords: quantum mechanical calculations ; substrate-enzyme interactions ; mutants ; functional role of amino acids ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The enzyme herpes simplex virus type 1 thymidine kinase (HSV1 TK) salvages thymidine into the DNA metabolism of the virus. In the active site, the thymine ring of the nucleoside binds in a pocket, formed by two residues, Tyr-172 and Met-128, in a sandwich-type orientation. To investigate the nature of the thymine-enzyme pocket interactions, we have carried out density functional theory calculations with gradient-corrected exchange-correlation functionals of models of the thymine-HSV1 TK adduct. Our calculations indicate that the role of Met-128 in the substrate fixation is purely steric and hydrophobic, while the substrate-Tyr-172 interaction is essentially electrostatic in nature. These findings are completely consistent with the available catalytic properties of mutants on the 128 position. Proteins 31:453-459, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
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  • 94
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    Electrostatics, allostery, and activity of the yeast chorismate mutase (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 445-452 
    Details
    ISSN: 0887-3585
    Keywords: chorismate mutase ; activity ; allosteric ; electrostatics ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The predicted active site of chorismate mutase of baker's yeast Saccharomyces cerevisiae has been studied by continuum electrostatics, molecular surface/volume calculations, and molecular modeling. Our study shows that despite being subject to an allosteric transition, the enzyme's active-site pocket neither decreased in volume nor deformed significantly in shape between the active R state and the inactive T state. We find that the polar atmosphere in the pocket is responsible for the enzyme's affinity. A single amino acid, Glu23, can adequately account for the atmospheric variation. This residue swings into the active-site pocket from the R state to the T state. In the R state, Glu23 on helix H2 doubly pairs with Arg204 and Lys208 of H11, which is packed against H2. In the T state, a slide occurs between H11 and H2 such that Glu23 can no longer interact with Lys208 and competes with Asp24 for interacting with Arg204. Consequently, Glu23 is found in the T state to couple with Arg157, an active-site residue critical to substrate binding. The tandem sliding of H11 in both monomers profoundly changes the interactions in the dimer interface. The loop between H11 and H12 demonstrates the largest conformational change. Hence, we establish a connection between the allosteric transition and the activity of the enzyme. The conformational change in the transition is suggested to propagate into the active-site pocket via a series of polar interactions that result in polarity reversal in the active-site pocket, which regulates the enzyme's activity. Proteins 31:445-452, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
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  • 95
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    Evidence of new cadmium binding sites in recombinant horse L-chain ferritin by anomalous Fourier difference map calculation (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 31 (1998), S. 477-485 
    Details
    ISSN: 0887-3585
    Keywords: X-ray structure ; L-chain apoferritin ; metal binding sites ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: We refined the structure of the tetragonal form of recombinant horse L-chain apoferritin to 2.0 Å and we compared it with that of the cubic form previously refined to the same resolution. The major differences between the two structures concern the cadmium ions bound to the residues E130 at the threefold axes of the molecule. Taking advantage of the significant anomalous signal (f′′ = 3.6 e-) of cadmium at 1.375 Å, the wavelength used here, we performed anomalous Fourier difference maps with the refined model phases. These maps reveal the positions of anomalous scatterers at different locations in the structure. Among these, some are found near residues that were known previously to bind metal ions, C48, E57, C126, D127, E130, and H132. But new cadmium binding sites are evidenced near residues E53, E56, E57, E60, and H114, which were suggested to be involved in the iron loading process. The quality of the anomalous Fourier difference map increases significantly with noncrystallographic symmetry map averaging. Such maps reveal density peaks that fit the positions of Met and Cys sulfur atoms, which are weak anomalous scatterers (f′′ = 0.44 e-). Proteins 31:477-485, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
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  • 96
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    The biology workbench - A seamless database and analysis environment for the biologist (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 32 (1998), S. 1-2 
    Details
    ISSN: 0887-3585
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: No abstract.
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  • 97
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    Calculation of HyHel10-lysozyme binding free energy changes: Effect of ten point mutations (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 39-48 
    Details
    ISSN: 0887-3585
    Keywords: antibody ; antigen ; electrostatics ; binding ; finite difference ; Poisson-Boltzmann ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The change in free energy of binding of hen egg white lysozyme (HEL) to the antibody HyHel-10 arising from ten point mutations in HEL (D101K, D101G, K96M, K97D, K97G, K97G, R21E, R21K, W62Y, and W63Y) was calculated using a combination of the finite difference Poisson-Boltzmann method for the electrostatic contribution, a solvent accessible surface area term for the non-polar contribution, and rotamer counting for the sidechain entropy contribution. Comparison of experimental and calculated results indicate that because of pKa shifts in some of the mutated residues, primarily those involving Aspartate or Glutamate, proton uptake or release occurs in binding. When this effect was incorporated into the binding free energy calculations, the agreement with experiment improved significantly, and resulted in a mean error of about 1.9 kcal/mole. Thus these calculations predict that there should be a significant pH dependence to the change in binding caused by these mutations. The other major contributions to binding energy changes comes from solvation and charge charge interactions, which tend to oppose each other. Smaller contributions come from nonpolar interactions and sidechain entropy changes. The structures of the HyHel-10-HEL complexes with mutant HEL were obtained by modeling, and the effect of the modeled structure on the calculations was also examined. “Knowledge based” modeling and automatic generation of models using molecular mechanics produced comparable results. Proteins 33:39-48, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 2 Ill.
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  • 98
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    Crystal structure of a complex formed between proteolytically-generated lactoferrin fragment and proteinase KAtomic coordinates have been deposited with the Protein Data Bank, Brookhaven National Laboratory. (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 30-38 
    Details
    ISSN: 0887-3585
    Keywords: lactoferrin ; proteinase K ; complex, hydrolysis ; structure ; inhibitor ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Lactoferrin is an iron binding glycoprotein with a molecular weight of 80 kDa. The molecule is divided into two lobes representing the N-terminal and C-terminal halves of the polypeptide chain, each containing an iron binding site. The serine proteinases such as trypsin, chymotrypsin, and pepsin hydrolyze lactoferrin into two unequal halves while proteinase K divides this protein into two equal halves. In the first step of hydrolysis by proteinase K, the C- and N-lobes, each having a molecular weight of approximately 40 kDa, are generated. In the next step, the lobes are further hydrolyzed into small molecular weight peptides. The proteinase K isolated from the hydrolyzed product does not show enzymatic activity suggesting that the enzyme is inhibited. Furthermore, the hydrolysis experiments on N-lobe and C-lobe showed that the inhibitory fragment came from the C-lobe. The purified lactoferrin fragment was found to be a decapeptide with an amino acid sequence of H2N-Val-Ala-Gln-Gly-Ala-Ala-Gly-Leu-Ala-COOH. The complex formed between proteinase K and lactoferrin fragment was crystallized by microdialysis. The crystals belonged to the monoclinic space group P21with cell dimensions a = 44.4 Å, b = 38.6 Å, c = 79.2 Å, β = 105.8o and Z = 2. The crystal structure has been determined at 2.4 Å resolution. It has been refined to an R factor of 0.163 for 9044 reflections. The Lf-fragment forms several intermolecular interactions with proteinase K. The Ser-224 Oγ and His-57 Nε2 move away to a distance of 3.68 Å in the complex. In the crystal structure, Gln-3I (I indicates inhibitor i.e., lactoferrin fragment) is involved in a direct intermolecular interaction with a symmetry related proteinase K molecule through a strong hydrogen bond with Asp-254. The mode of intermolecular interactions in the complex conformational features of the enzyme and placement of the fragment with respect to the enzyme resemble with the molecular complex of proteinase K with its natural inhibitor PKI3 from wheat. Proteins 33:30-38, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 8 Ill.
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  • 99
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    Electronic Resource
    Dictionary of recurrent domains in protein structures (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 88-96 
    Details
    ISSN: 0887-3585
    Keywords: fold classification ; substructures ; Dali ; protein families ; structural similarity ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The rapid growth in the number of experimentally determined three-dimensional protein structures has sharpened the need for comprehensive and up-to-date surveys of known structures. Classic work on protein structure classification has made it clear that a structural survey is best carried out at the level of domains, i.e., substructures that recur in evolution as functional units in different protein contexts. We present a method for automated domain identification from protein structure atomic coordinates based on quantitative measures of compactness and, as the new element, recurrence. Compactness criteria are used to recursively divide a protein into a series of successively smaller and smaller substructures. Recurrence criteria are used to select an optimal size level of these substructures, so that many of the chosen substructures are common to different proteins at a high level of statistical significance. The joint application of these criteria automatically yields consistent domain definitions between remote homologs, a result difficult to achieve using compactness criteria alone. The method is applied to a representative set of 1,137 sequence-unique protein families covering 6,500 known structures. Clustering of the resulting set of domains (substructures) yields 594 distinct fold classes (types of substructures). The Dali Domain Dictionary (http://www.embl-ebi.ac.uk/dali) not only provides a global structural classification, but also a comprehensive description of families of protein sequences grouped around representative proteins of known structure. The classification will be continuously updated and can serve as a basis for improving our understanding of protein evolution and function and for evolving optimal strategies to complete the map of all natural protein structures. Proteins 33:88-96, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
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  • 100
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    Crystal structures of HLA-A*0201 complexed with antigenic peptides with either the amino- or carboxyl-terminal group substituted by a methyl group (1998)
    New York, NY : Wiley-Blackwell
    Proteins: Structure, Function, and Genetics 33 (1998), S. 97-106 
    Details
    ISSN: 0887-3585
    Keywords: antigenic peptides ; class I MHC molecules ; HLA-A2 complexes ; hydrogen bonds ; protein structure ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The crystal structures of class I major histocompatibility complex (MHC) molecules complexed with antigenic peptides revealed a network of hydrogen bonds between the charged amino- and carboxyl-termini of the peptides and conserved MHC residues at both ends of the peptide binding site. These interactions were shown to contribute substantially to the stability of class I MHC/peptide complexes by thermal denaturation studies using synthetic peptides in which either the amino- or carboxyl-terminal group is substituted by a methyl group. Here we report crystal structures of HLA-A*0201 complexed with these terminally modified synthetic peptides showing that they adopt the same bound conformation as antigenic peptides. A number of variations in peptide conformation were observed for the terminally modified peptides, including in one case, a large conformational difference in four central peptide residues that is apparently caused by the lattice contact. This is reminiscent of the way binding a T-cell receptor changed the conformation of central residues of an MHC-bound peptide. The structures determined identify which conserved hydrogen bonds are eliminated in terminally substituted peptides and suggest an increased energetic importance of the interactions at the peptide termini for MHC-peptide stability. Proteins 33:97-106, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 4 Ill.
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