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  • 101
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 333-338 
    ISSN: 0192-253X
    Keywords: Cell migration ; Aphidicolin ; Blastula-Gastrula ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Inhibition of DNA replication by aphidicolin in the chick morula interferes with its progression to a normal blastula and prevents induction of the first morphogenetic cell movements of primitive streak formation. Embryos in aphidicolin synthesize some polypeptides typical of blastula but do not display all the characteristic features of morula to blastula transition. Inhibition of DNA replication inteferes with the sequential synthesis of maternally coded polypeptides and with the activation of the embryonic genome in the chick embryo.
    Additional Material: 4 Ill.
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  • 102
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 345-345 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 103
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 347-347 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 104
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989) 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 105
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 339-344 
    ISSN: 0192-253X
    Keywords: Delayed processing ; Splicing ; Transcription ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: This study deals with the pattern of developmental expression of the catalase gene in mice. We have used a mouse catalase 2 kb cDNA (pMCT-1) and its 1.4 kb 5′ fragment as probes to characterize the transcripts during embryonic development and differentiation. Total RNA was isolated from 8 days postconceptus (p.c.) whole embryos and from livers and carcasses of 13, 15, and 18 day p.c. embryos as well as from the livers of newborn and adult mice of the S.W. strain. The RNA was applied on slot blots, and run on agarose gels to generate northern blots. Blots were hybridized with the 32P-labeled cDNA probe under different stringency conditions. Autoradiograms were scanned with a densitometer to quantify relative hybridization signals of RNA samples obtained from two or three individual mice representing each stage of development.The catalase transcript is detectable as early as 8 days p.c. with the beginning of somite formation. At this stage, it is primarily in the form of a 12.2 kb transcript. One additional band (2.4 kb) is also apparent at this stage although at a very low intensity. The intensity of the two bands increases with development, particularly during 13-18 days p.c. in liver and carcass. The 2.4 kb RNA band increases sharply from day 8 through 13, 15, and 18 days p.c. and is confined primarily to the liver. Interestingly, only the 2.4 kb RNA band is seen at and after birth. The 2.4 kb RNA is the known mature message of the catalase gene in mice. The presence of large catalase-specific RNA species (seen during development in utero only) is interpreted as the primary transcript of this gene. The complete and efficient processing of this primary transcript takes place only after birth and primarily in the liver, which may be related to the physiological role of this enzyme in oxygen metabolism, particularly stressful superoxides, expected with independent respiration. At a lower stringency wash of the northern blots, a 9.5 kb RNA was seen during a narrow window of in utero development. This 9.5 kb band may represent an uncharacterized catalase-related gene with a possible role in development and differentiation.
    Additional Material: 4 Ill.
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  • 106
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 349-355 
    ISSN: 0192-253X
    Keywords: SV40 promoter ; Expression vector ; Drug resistance ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have previously demonstrated systemic resistance to methotrexate (MTX) in transgenic mice carrying a foreign, mutant dihydrofolate reductase (DHFR, E.C. 1.5.1.3) gene. The new gene was introduced as a cDNA cloned into an expression vector driven by the simian virus 40 (SV40) early promoter. Previous physiologic studies suggested that transgenic mice tolerated drug doses invariably lethal to controls on the basis of gastrointestinal (GI) resistance to MTX. In the present study we evaluated foreign gene expression at the RNA level in the three major sites of MTX toxicity: intestine, liver, and bone marrow.The transgene was transcriptionally active in small bowel, and levels of expression were high in animals tolerating the largest doses of MTX. The gene was also expressed in the liver in some pedigrees, but was not detected in hemopoietic tissues of any of the pedigrees tested. Our studies correlate the site of expression of a drug resistant dhfr gene with an altered physiologic response to MTX, and demonstrate that transgenic mice can be used as a test system for expression of genes considered for use in somatic gene therapy.
    Additional Material: 8 Ill.
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  • 107
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 356-364 
    ISSN: 0192-253X
    Keywords: Glucose intolerance ; Insulin resistance ; Diabetes mellitus ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We are investigating human insulin gene expression in transgenic mice. An 8.8 kilobase (kb) human genomic DNA fragment, including the insulin gene (1.4 kb) and 2 kb of 5′ human flanking sequences, was introduced into mouse embryos by pronuclear microinjection. Two lines of transgenic mice have been established, both of which carry the intact human gene in multiple copies. Animals from both lines have significantly higher insulin levels than control mice, and the degree of hyperinsulinemia shows a positive correlation with human gene copy number in the two lines. Expression of the human gene is confirmed by the detection of human C-peptide in plasma. Tissue specificity of expression is maintained, with human insulin mRNA detectable only in the pancreas. The transgenics maintain normal fasting blood glucose in spite of their high insulin levels, but preliminary studies show them to be glucose intolerant when given a glucose load. These mice provide a model system for further studies on the regulation of insulin gene expression and on the effects of chronic hyperinsulinemia on glucose homeostasis.
    Additional Material: 6 Ill.
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  • 108
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989) 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 109
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 411-411 
    ISSN: 0192-253X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 110
    Electronic Resource
    Electronic Resource
    Chichester [u.a.] : Wiley-Blackwell
    Developmental Genetics 10 (1989), S. 402-410 
    ISSN: 0192-253X
    Keywords: F9 ECC ; Aggregates ; Embryoid bodies ; Endoderm ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: To study the relationship between compaction and differentiation in aggregates of F9 embryonal carcinoma cells, a subline was developed which grows mostly uncompacted in monolayer culture in medium containing a low concentration of calcium (about 0.05 mM). When these cells were trypsinized and cultured in suspension in the same medium, they formed loose, open aggregates, which failed to differentiate into embryoid bodies after exposure to 10 nM retinoic acid, confirming the requirement of compaction for differentiation. If, after culture for 3 days, the uncompacted F9 aggregates were exposed to additional calcium (4 mM), all compacted within an hour. The number of days necessary for aggregates to acquire this ability to compact rapidly was reduced if the monolayer of cells from which the aggregates were derived had been exposed to additional calcium to cause compaction for several days prior to trypsinization and aggregation. Next, treatment of the compacted F9 aggregates with 10 nM retinoic acid was followed by differentiation into embryoid bodies. The number of days required for this was also reduced if the aggregates were formed from previously compacted cells, presumably because compaction of the aggregates occured sooner.The acceleration in compaction and differentiation in aggregates formed from previously compacted cells suggests that some of the proteins important for compaction, which are synthesized in a monolayer of compacted cells, persist through trypsinization and are carried over from monolayer to aggregates. Alternatively, an inhibitor of compaction is decreased in the compacted monolayer. Thus, the process of compaction in its entirety, including its relationship to subsequent differentiation, cannot be studied in aggregates formed from F9 cells grown as usual in the compacted state in monolayer culture. This work provides an alternative system in which aggregation, compaction, and differentiation of F9 cells can be made to occur in stepwise fashion and can be examined separately.
    Additional Material: 6 Ill.
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  • 111
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. S339 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 112
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. S505 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 113
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. S303 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 114
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 115
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 116
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 1-10 
    ISSN: 0749-503X
    Keywords: Yeast ; genome size ; orthogonal-field-alternation gel electrophoresis ; mitochondrial DNA ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Using an improved procedure of pulsed field gel electrophoresis, yeast chromosomes were separated over a wide range of molecular size (250-4000 kbp) on single gels. The chromosomal DNA patterns of all the species belonging to the genus Kluyveromyces were examined. Within the species K. marxianus, the varieties lactis, drosophilarum and vanudenii showed closely related patterns; very different from them, the varieties bulgaricus and marxianus were related to each other, forming a distinct group; the strains commonly called ‘K. lactis’ and ‘K. fragilis’ were unambiguously different from each other in chromosome patterns. These differences were correlated with the presence of characteristic repetitive sequence elements in the mitochondrial DNA of the former group and not in the latter. Analysis of Candida macedoniensis, which had been considered to be an anamorph of K. marxianus var. marxianus, showed that these two yeast species were indeed similar in chromosome patterns and in mitochondrial DNA restriction patterns.
    Additional Material: 3 Ill.
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  • 117
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 25-33 
    ISSN: 0749-503X
    Keywords: Secretion ; Saccharomyces cerevisiae ; Golgi apparatus ; protein targetting ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The KEX2 protease (product of the KEX2 gene) functions late in the secretory pathway of Saccharomyces cerevisiae by cleaving the polypeptide chains of prepro-killer toxin and prepro-α-factor at paired basic amino acid residues. The intracellular vesicles containing KEX2 protease sedimented in density gradients to a position distinct from those containing mannosyltransferase I (product of the MNN1 gene), a marker enzyme for the Golgi complex. The recovery of intact compartments containing these enzymes approached 80% after sedimentation. We propose that the KEX2 protease and mannosyltransferase I reside within distinct compartments.
    Additional Material: 3 Ill.
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  • 118
    ISSN: 0749-503X
    Keywords: Yeast ; α-glucosidase ; nucleotide sequence ; expression ; proteinase ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Two α-glucosidase (maltase) genes, designated GLUCPI and GLUCPII, have been cloned from an industrial strain of baker's yeast (Saccharomyces cerevisiae) by complementation of a maltase-negative mutant strain. The different genes were identified according to their alternatively expressed isoenzymes PI and PII in transformants after isoelectric focusing and activity staining in separated cell lysates. The gene encoding α-glucosidase PI (GLUCPI), which was not present in laboratory strains of S. carlsbergensis with a defined MAL1, 2, 3, 4 or 6 locus, was sequenced and compared with the recently published MAL6S gene. This comparison revealed single amino acid deviations at three positions in the predicted polypeptide sequence. In addition, the divergent promoter region of GLUCPI differed from MAL6S by a triple repeated 147-bp DNA segment. Maltose induction and glucose repression of α-glucosidase PI were not affected by the deletion of the repeated DNA segment. However, the absolute expression of α-glucosidase PI increased two- to four-fold. In addition, a two-fold increase in the maltase synthesis occurred when the cloned positive regulator gene MAL2-8cp was on the same plasmid. Furthermore, stability of the α-glucosidase in cultures in the stationary growth phase was greatly enhanced using a host strain lacking the proteinases A and B and the carboxypeptidases Y and S. Promoter trimming, MAL2-8cp stimulation and the use of a host strain deficient in four vacuolar proteinases resulted in α-glucosidase PI expression of about 13% of the soluble protein.
    Additional Material: 7 Ill.
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  • 119
    ISSN: 0749-503X
    Keywords: TRP1 ; histone H3 ; histone H4 ; pyrophosphatase ; Kluyveromyces ; yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The TRP1 gene of the yeast Kluyveromyces lactis has been cloned from a genomic library by complementation of the Saccharomyces cerevisiae trpl-289 mutation. The gene was located within the clone by transposon mutagenesis and the coding region identified by DNA sequencing. This has indicated that K. lactis TRP1 encodes a 210-amino acid polypeptide which shows 53% identity to the homologous S. cerevisiae protein. The K. lactis TRP1 gene has been disrupted by substituting the S. cerevisiae URA3 gene for a large part of the TRP1 coding sequence. Replacement of the chromosomal TRP1 locus with this construction has enabled the production of non-reverting trp1- strains of K. lactis, while a genetic analysis of the disrupted allele confirmed that the TRP1 gene had been cloned. DNA sequencing has also shown that the K. lactis TRP1 sequences is flanked by genes encoding inorganic pyrophosphatase and histone H3, which we have designated IPP and HHT1 respectively. Hybridization studies have shown that in common with S. cerevisiae, K. lactis has two copies of the histone H3 gene. Each H3 gene is closely linked to a gene encoding histone H4 and in both yeast species the IPP gene is tightly linked to one of the histone gene pairs.
    Additional Material: 7 Ill.
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  • 120
    ISSN: 0749-503X
    Keywords: Yeast ; protein ; extract ; trichloroacetic acid ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Methods currently used for the extraction of proteins from yeast involve relatively long time periods between sampling cells from a culture and analysis of their proteins by polyacrylamide gel electrophoresis-sodium dodecylsulphate. Often it is desirable to inactivate cellular metabolism rapidly after sampling and here we show that trichloroacetic acid precipitation techniques, often used for rapid extraction and inactivation of proteins from higher eukaryotes, can be adapted for use with organisms which have cell walls.
    Additional Material: 1 Ill.
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  • 121
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 122
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 123
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 55-72 
    ISSN: 0749-503X
    Keywords: Gene disruption ; genetic mapping ; nonsense suppression ; multibudded phenotype ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A newly isolated gene, ESS1, was shown to encode a protein required for vegetative growth in Saccharomyces cerevisiae. The nucleotide sequence of ESS1 revealed a 172 amino acid open reading frame predicting a highly basic, 19·5 kilodalton product. Although the gene was isolated by cross-hybridization with the vertebrate v-sis oncogene, the primary amino acid sequence bears only a slight resemblance to the p28sis protein. ESS1 was shown to be single copy in the yeast genome and transcriptionally active during logarithmic growth. It is located on the right arm of chromosome X, 6 centimorgans distal to ilv3. The genetic map location indicates it is not allelic to any previously characterized mutation in this organism. Both inactivation of ESS1 by gene disruption and overexpression by fusion to a heterologous promoter were detrimental to growth in both haploid and diploid cell types. Under non-permissive conditions, the terminal phenotype of strains containing a suppressible amber mutation within ESS1 was one of aberrant multibudded structures. Examination of this morphology indicates that loss of ESS1 function may lead to a defect in cytokinesis or cell separation.
    Additional Material: 7 Ill.
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  • 124
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 73-77 
    ISSN: 0749-503X
    Keywords: vandate ; mitochondria ; H+ ATPase ; S. cerevisiae ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The effects of vandate on mitochondrial respiration and H+ ATPase activity in Saccharomyces cerevisiae were studied. A 50% inhibition of oxygen uptake in isolated mitochondria was produced by 4·4 mM-V2O5. Activity of H+ ATPase in whole mitochondria was inhibited by 50% by 5·5 μM-V2O5, in submitochondrial particles by 55 μM-V2O5; and in the chloroform-released H+ ATPase by 0·5 mM-V2O5. Vandate was also found to relieve growth inhibition caused by the mitochondrial H+ ATPase inhibitors NN′-decyclohexylcarbodiimide and oligomycin. These results imply that vanadate could affect mitochondrial respiration by interacting with the H+ ATPase in S. cerevisiae.
    Additional Material: 1 Ill.
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  • 125
    ISSN: 0749-503X
    Keywords: CDC33 ; cell division cycle ; cyclic AMP ; start gene ; yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The CDC33 gene of Saccharomyces cerevisiae belongs to the class II ‘START’ genes. Its product is required for the initiation of a new cell division cycle (Hartwell, 1974). Many results suggest that the cAMP signalling pathway is one of the major controlling elements of ‘START’. Components of this pathway are encoded by class II ‘START’ genes. The aim of the present study is to determine whether or not the CDC33 gene interferes with the cAMP signalling pathway. We report here the molecular cloning of the CDC33 gene by complementation of the cdc33-1 thermosensitive mutant. The identity of the cloned gene is confirmed by site-specific reintegration and segregation analysis. This gene is transcribed into a 900-nucleotides mRNA and appears to be relatively abundant in the cell. We also show that the CDC33 gene product is essential for sporulation. cdc33-1 mutant cells are able to enter into the resting state. The cAMP intracellular pool is not modified when the cdc33-1 mutant is shifted to the restrictive temperature. The cdc33-1 mutation is not suppressed by other known elements of the cAMP cascade. All these results suggest that the CDC33 ‘START’ gene does not interfere with the cAMP signalling pathway which controls cell division.
    Additional Material: 6 Ill.
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  • 126
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    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 91-98 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; respiratory-deficient mutants ; increased gene expression ; mRNA level ; human lysozyme ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Respiratory-deficient mutants (rho- cells) of Saccharomyces cerevisiae produced about 10 times as much human(h-) lysozyme as did wild-type strains (rho+ cells) when the GAL10 promoter was used in an expression plasmid with the h-lysozyme gene. Introduction of intact mitochondria into the rho- cells resulted in a significant decrease in the production of h-lysozyme, indicating that the rho- mutation increased the expression of the h-lysozyme gene. The copy number of the expression plasmid was not responsible for the increased expression. The level of h-lysozyme mRNA in the rho- cells was also much higher than that in the rho+ cells especially at the stationary phase. The increased expression of the h-lysozyme gene was also observed when a glyceraldehyde-3-phosphate dehydrogenase gene promoter and the PHO5 promoter were used in the expression plasmid. The rho- mutation also increased the expression of the PHO5 gene under the control of the HIS5 promoter in a plasmid and the ACT1 gene in the yeast chromosome, but did not increase the expression of the ribosomal RNA gene. In contrast to the rho- mutants, pet mutants did not show higher gene expression compared with wild-type strains.
    Additional Material: 3 Ill.
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  • 127
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 107-115 
    ISSN: 0749-503X
    Keywords: Pichia pastoris ; glycoproteins ; invertase ; oligosaccharides ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The secreted glycoproteins of Pichia pastoris contain more than 35% of their N-linked oligosaccharides as structures smaller than Man14GlcNAc2 (Man = mannose; GlcNAc = N-acetylglucosamine). On heterologous invertase produced in P. pastoris, approximately 85% of the oligosaccharides are in the size range Man8-14GlcNAc2. The structures appear to contain α-linked mannose. In addition, one-third of the structures contain net negative charge and can be radio-labelled in vivo with 32P. The largest oligosaccharides isolated from P. pastoris are significantly shorter than the hypermannosylated structures typical of S. cerevisiae, indicating that the factors which influence the processing of N-linked oligosaccharides in P. pastoris are different from those which influence processing in S. cerevisiae. The smaller N-linked oligosaccharides synthesized by P. pastoris resemble high-mannose oligosaccharides synthesized by animal cells, and this finding increases the utility of P. pastoris as a host for the production of heterologous glycoproteins.
    Additional Material: 3 Ill.
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  • 128
    ISSN: 0749-503X
    Keywords: Kluyveromyces lactis ; mitochondrial respiration ; erythromycin ; sugar metabolism ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The analysis of five independent isolates of Kluyveromyces lactis shows that CBS 2359, CBS 683 and CBS 4574 could grow in the presence of mitochondrial inhibitors (antimycin A, oligomycin or erythromycin) and that CBS 2360 and CBS 141 were unable to grow in the presence of drugs. The resistant growth was observed only on glucose and not on other fermentable carbon sources (galactose, lactose).The phenotype ‘growth on glucose in the presence of mitochondrial inhibitors’ was called Rag+. This phenotype was found to be controlled by two unlinked nuclear genes: RAG1 and RAG2. Either of their recessive alleles, rag1 and rag2, led to the Rag- phenotype (i.e. the failure of growth on glucose in the presence of antimitochondrial drugs).Rag- strains represent the case in which fermentative growth becomes absolutely dependent on the functioning of the normal respiratory chain.
    Additional Material: 2 Ill.
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  • 129
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 130
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 131
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 131-139 
    ISSN: 0749-503X
    Keywords: Yeast ; γ-irradiation ; post-irradiation recovery ; radiosensitive mutants ; DNA double-strand break repair ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: G1 cells of the diploid yeast Saccharomyces cerevisiae are known to be capable of a slow repair of DNA double-strand breaks (DSB) during holding the cells in a non-nutrient medium (Luchnik et al., 1977; Frankenberg-Schwager et al., 1980). In the present paper, S. cerevisiae cells γ-irradiated in the G1 phase of the cell cycle are shown to be capable of fast repair of DNA DSB; this process is completed within 30-40 min, of holding the cells in water at 28°C. For this reason, the kinetics of DNA DSB repair during holding the cells in a non-nutrient medium are biphasic, i.e., the first ‘fast’ phase is completed within 30-40 min, whereas the second, ‘slow’ phase is completed within 48 h. Mututions rad51, rad52, rad54 and rad55 inhibit the fast repair of DNA DSB, whereas mutations rad50, rad53 and rad57 do not significantly influence this process.It has been shown that the observed fast and slow repair of DNA DSB in the G1 diploid cells of S. cerevisiae are separate pathways of DNA DSB repair in yeast.
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  • 132
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    Yeast 5 (1989), S. 117-129 
    ISSN: 0749-503X
    Keywords: Cell cycle ; synchronization ; DNA replication ; killer ; in vitro replication ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A detailed characterization of the mak 1-3 mutation of Saccharomyces cerevisiae has been made possible by modifying its genetic background. The mak1-3 mutation, which confers temperature sensitivity for growth, was originally identified as one of four mak1 mutations (Wickner and Leibowitz, 1976). Mak1-1, 1-2 and 1-4 mutants are deficient in DNA topoisomerase I activity and thus have been renamed ‘top1’ (Thrash et al., 1984). Studies presented here show that the map position of MAK1-3 on chromosome XVI distinguishes it from TOP1 which maps on chromosome XV (Wickner and Leibowitz, 1976). An investigation of in vivo macromolecular synthesis in the mak1-3 mutant shows that it is deficient in DNA replication at the restrictive temperature. Experiments in which DNA synthesis was measured in synchronized cell populations indicate that the mak1-3 mutant is deficient in the initiation step of DNA synthesis. Furthermore, crude extracts from the mak1-3 mutant cells support temperature-sensitive in vitro DNA synthesis on yeast chromosomal DNA replication origin containing plasmid pARS1, suggesting that the MAK1 gene product is directly required for in vitro DNA replication. The conclusion that mak1-3 is a newly identified DNA replication mutation is based on the observations that it (1) complements all DNA synthesis mutants examined, (2) maps to a previously undetected chromosomal location and (3) has a distinct terminal morphology. In light of these distinctions and of the role mak1-3 plays in DNA replication, it has been renamed ‘dnal’.
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  • 133
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    Yeast 5 (1989), S. 149-158 
    ISSN: 0749-503X
    Keywords: Superkiller ; double-stranded RNA ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The yeast chromosomal genes SK12, SK13, SK14, SK16, SK17 and SK18 repress the replication of double-stranded RNA viruses, protecting the host from the otherwise lethal effects of the virus. We cloned and sequenced the SK13 gene and found that it encodes a 163 kDa protein including a typical nuclear localization signal. Cell fractionation experiments show that the SK13 gene product is indeed tightly associated with nuclei and that the putative nuclear localization sequence directs β-galactosidase into the nucleus. However, fusion of a part of the SK13 protein lacking this signal with β-galactosidase directs β-galactosidase into the nucleus, suggesting the presence of a second nuclear localization signal. The SK13 gene is only essential in the presence of an M double-stranded RNA virus.
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  • 134
    ISSN: 0749-503X
    Keywords: Yeast ; cytochrome P-450 ; UV and X-ray irradiation ; oxidative damage ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Cytochrome P-450 was induced both in the diploid wild-type D7 strain and in two isogenic DNA-repair-deficient strains (rad3 and rad56) of Saccharomyces cerevisiae following UV- and X-irradiation. The induction occurred only in logarithmic growth phase cells and it was transient showing a peak 3 h after irradiation. The maximal amount of cytochrome P-450 was directly proportional to the radiation does applied. Under the same experimental conditions an increase of the catalase activity was also observed, suggesting that activated oxygen species produced by irradiation might be implicated in the induction of both enzymes. The sensitivity to H2O2 of cells containing high cytochrome P-450 levels was enhanced when this enzyme was specifically inhibited by tetrahydrofuran and metyrapone. This supports the hypothesis that cytochrome P-450, as well as catalase, might be involved in cell protection against oxidative damage.
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  • 135
    ISSN: 0749-503X
    Keywords: Crabtree effect ; sugar transport ; growth kinetics ; yeast ; chemostat ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The glucose transport capacity of Saccharomyces cerevisiae CBS 8066 was studied in aerobic glucose-limited chemostat cultures. Two different transport systems were encountered with affinity constants of 1 and 20 mM, respectively. The capacity of these carriers (Vmax) was dependent on the dilution rate and the residual glucose concentration in the culture. From the residual glucose concentration in the fermenter and the kinetic constants of glucose transport, their in situ contribution to glucose consumption was determined. The sum of these calculated in situ transport rates correlated well with the observed rate of glucose consumption of the culture.The growth kinetics of S. cerevisiae CBS 8066 in glucose-limited cultures were rather perculiar. At low dilution rates, at which glucose was completely respired, the glucose concentration in the fermenter was constant at 110 μM, independent of the glucose concentration in the reservoir. At high dilution rates, characterized by the occurrence of both respiration and alcoholic fermentation, the residual substrate concentration followed Monod kinetics. In this case, however, the overall affinity constant was dependent on the reservoir glucose concentration.
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  • 136
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    Yeast 5 (1989), S. 167-177 
    ISSN: 0749-503X
    Keywords: Methylotrophic yeasts ; alcohol oxidase ; Pichia pastoris ; genome evolution ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In methylotrophic yeasts, alcohol oxidase is the first enzyme in the methanol-utilization pathway. The genome of one such yeast, Pichia pastoris, contains two alcohol oxidase genes, AOX1 and AOX2. Sequence analysis indicated that each gene encodes a similar protein of 663 amino acids. The protein-coding regions of the genes were 92% and 97% homologous at the nucleotide and predicted amino acid sequence levels, respectively. In contrast to homology observed within the protein-coding portions of the AOX genes, no homology was found in either the 5′ or 3′ non-coding regions. Although alcohol oxidase is found in peroxisomes of P. pastoris, the AOX amino acid sequences did not contain a peptide sequence similar to the peroxisomal transport sequence found at the C-terminus of some peroxisomally located proteins in higher eukaryotes.
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  • 137
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    Yeast 5 (1989), S. 179-186 
    ISSN: 0749-503X
    Keywords: Pichia pinus ; yeast ; mutants ; ethanol metabolism ; methanol oxidation ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A collection of mutants of Pichia pinus which are unable to grow on ethanol but retain the ability to grow on glucose and methanol, was obtained. Genetic and biochemical analysis of these strains revealed mutations in seven nuclear genes affecting activities of isocitrate lyase (icl1), malate synthase (mls1), phosphoenolpyruvate carboxykinase (pck1), ‘malic’ enzyme (mdd1) and acetyl-CoA synthetase (acs1, acs2 and acs3). All mutations except acs1-acs3 have no effect on the activities of other enzymes involved in C2 metabolism. Mutations acs1, acs2 and acs3 have a pleiotropic action, leading to partial reduction in activities of isocitrate lyase and malate synthase. Ethanol-induced repression of the synthesis of the methanol oxidative enzymes, alcohol oxidase and catalase, is not impaired in these seven mutant classes. On the other hand, C2 compound-induced inactivation of alcohol oxidase and catalase is impaired in mutants acs1, acs2, acs3 and icl1. It was suggested that glyoxylate and acetate (or acetate precursors) act as low molecular weight effectors, ‘switching on’ inactivation and repression, respectively, of alcohol oxidase and catalase in the medium containing ethanol or acetate.
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  • 138
    ISSN: 0749-503X
    Keywords: Yeast ; messenger RNA ; translation ; codon bias ; RNA secondary-structure ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The effects of poor codon bias and secondary structure formation upon the translation of the pyruvate kinase (PYK1) mRNA have been investigated in Saccharomyces cerevisiae. Following insertion mutagenesis at the 5′-end of the PYK1 coding region, the gene was transformed into yeast, and translation assessed directly in vivo by determining the distribution of the modified PYK1 mRNAs across polysomes fractionated by sucrose density gradient centrifugation. The chromosomally-encoded (wild-type) PYK1 mRNA, and the actin, ribosomal protein L3 and glyceraldehyde-3-phosphate dehydrogenase mRNAs were used to control for minor differences between polysome preparations. An insertion containing 13 non-preferred codons at the 5′-end of the coding region was found to have no significant effect upon PYK1 mRNA translation. In contrast, translation was inhibited by an insertion which increased the formation of secondary structures at the 5′-end of the mRNA (overall ΔG = -36·6 kcal/mol). Control insertions were also analysed to exclude the possibility that alterations to the amino acid sequence of pyruvate kinase affect the translation of its mRNA. These insertions, which introduced preferred codons or restored wild-type levels of secondary structure formation, did not significantly influence PYK1 mRNA translation.
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  • 139
    ISSN: 0749-503X
    Keywords: Kluyveromyces lactis ; lactose-fermenting yeast ; cytochromes ; mitochondrial genes ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The apocytochrome b genes from two strains of the yeast Kluyveromyces lactis, have been isolated and sequenced. The coding sequences in strains WM27 (NRRL Y-17066) and WM37 (NRRL Y-1140) were identical but the upstream noncoding regions were slightly different. The sequences demonstrated the presence of a continuous open reading frame with no introns. The amino acid sequence, derived from the coding strand, showed 82% homology to the apocytochrome b of Saccharomyces cerevisiae strain D273-10B and only 58% homology to the protein from Schizosaccharomyces pombe strain 50. CUN and CGN codon families were absent from the K. lactis gene. Codon usage was very similar to that of other mitochondrial genomes with mostly U or A in the third position. There were two unusual features. All threonines were coded by ACA(U) and all arginines by AGA.
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  • 140
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    Yeast 5 (1989), S. 199-207 
    ISSN: 0749-503X
    Keywords: L-Leucine ; amino-acid transport ; Schizosaccharomyces pombe ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Transport of L-leucine into Schizosaccharomyces pombe cells from the stationary phase of growth (after preincubation for 60 min with 1% glucose) proceeds uphill, practically unidirectionally, and is mediated by at least two systems: a high-affinity system with a KT of 0·045 mmol 1-1 and Jmax of 3·3 nmol min-1 (mg dry weight)-1 and a low-affinity system with a KT of 1·25 mmol 1-1 and Jmax of 16·0 nmol min-1 (mg dry weight)-1. The high-affinity system has a pH optimum at 3.2, the accumulation ratio is highest at a cell density of 2-4 mg dry weight per ml and decreases with increasing leucine concentration. Transport of leucine by the high-affinity system is strongly inhibited by proton conductors, ammonium ions and by most amino acids, but only L-phenylalanine, L-isoleucine, L-valine and L-cysteine behave as fully competitive inhibitors. Systems of L-leucine transport in S. pombe are not constitutive. Transport activity appears only after preincubation of cells with a suitable source of energy. If cycloheximide is added during preincubation with glucose, no transport systems for leucine are synthesized. After removal of glucose, the activity of transport systems decays with a half-time of about 20 min. The presence of cyclic AMP increases the initial rate of leucine uptake only in cells preincubated with glucose and in the absence of cycloheximide.
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  • 141
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    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 142
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    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 143
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    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 144
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    Yeast 5 (1989), S. 219-238 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 145
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    Yeast 5 (1989), S. 239-257 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Additional Material: 7 Ill.
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  • 146
    ISSN: 0749-503X
    Keywords: DNA sequence ; gene function ; ORF ; S. cerevisiae ; transposon ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The development of mega-sequencing techniques requires new methods for global functional analysis of cloned DNA fragments. We have developed a mini-Mu transposon adapted to yeast cloned DNA fragment analysis. This transposon allows us to do the following in a single construction: (i) to probe yeast cells for the presence of expressed open reading frames (ORFs) in the cloned DNA fragment; (ii) to localize these ORFs in the fragment and determine their transcription orientation; (iii) to use β-galactosidase protein fusions to study regulation of these ORFs; and (iv) to disrupt the corresponding chromosomal genes.On a 5-kb yeast DNA sequence, we have verified the reliability of this new tool by comparing the data obtained with the mini-Mu transposon to those obtained by classical methods.This transposon should be of immediate use in the yeast genome sequencing programme.
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  • 147
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    Yeast 5 (1989), S. 285-290 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; glycolysis ; metabolic flux ; ethanol production ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Eight different enzyymes for glycolysis and alcoholic fermentation were overproduced in a common Saccharomyces cerevisiae strain by placing their genes on multicopy vectors. The specific enzyme activities were increased between 3·7-and 13·9-fold above the wild-type level. The overproduction of the different glycolytic enzymes had no effect on the rate of ethanol formation, even with those enzymes that catalyse irreversible steps: hexokinase, phosphofructokinase and pyruvate kinase. Also the simultaneous increase in the activities of pairs of enzymes such as pyruvate kinase and phosphofructokinase or pyruvate decarboxylase and alcohol dehyrogenase, did not increase the rate of ethanol production. The levels of key glycolytic metabolites were also normal, compared to the reference strain.
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  • 148
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    Yeast 5 (1989), S. 271-284 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have devised a genetic screen to identify trans-acting factors involved in chromosome transmission in yeast. This approach was designed to potentially identify a subset of genes encoding proteins that interact with centromere DNA. It has been shown that mutations in yeast centromere DNA cause aberrant chromosome segregation during mitosis and meiosis. We reasoned that the function of an altered centromere should be particularly sensitive to changes in factors with which it interacts. We constructed a disomic strain containing one copy of chromosome III with a wild-type centromere and one copy of chromosome III bearing the SUP11 gene and a mutant CEN3. This strain forms white colonies with red sectors due to nondisjunction of the chromosome bearing the mutant centromere. After mutagenesis we picked colonies that exhibited increased nondisjunction of the mutant chromosome as evidenced by increased red-white sectoring. Using this approach, we have isolated three trans-acting chromosome nondisjunction (cnd) mutants that are defective in maintaining chromosomes during mitosis in yeast.
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  • 149
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    Yeast 5 (1989), S. 291-298 
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; translation ; protein synthesis ; mRNA stability ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have characterized a nonsense mutation in the ADR1 gene that identifies the translational start of the ADR1 protein. The ADR1 gene of Saccharomyces cerevisiae is required for synthesis of the glucose-repressible alcohol dehydrogenase (ADH2). The adr1-1 mutation, which inhibits ADH2 expression, was identified as a C to G transversion at base pair +32. This alteration would result in a UGA nonsense codon in place of a serine codon that would lead to termination of the ADR1 polypeptide after the 10th amino acid. The effect of the adr1-1 mutation was partially reversed by UGA-tRNA suppressors, indicating that the adr1-1 mutation affects ADR1 expression at the translational level. These observations establish that the first available AUG in the ADR1 sequence is used as the translational start site of ADR1. Tyrosine or leucine UGA-tRNA-suppressors resulted in levels of adr1-1 activity similar to that found for a serine UGA-tRNA-suppressor, suggesting that serine residue-11 is not essential to ADR1 function. Northern analyses showed that the 5·1 kb ADR1 mRNA was two- to three-fold more abundant when isolated from a strain carrying the ADR1 allele than from an isogenic strain containing the adr1-1 allele. These data confirm that the 5·1 kb mRNA is the ADR1 mRNA and suggest that inhibition of adr1-1 mRNA translation results in more rapid degradation of the adr1-1 mRNA.
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  • 150
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    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 151
    ISSN: 0749-503X
    Keywords: Heterologous gene expression ; protein secretion ; S. cerevisiae ; glycosylation ; cellulases ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The Clostridium thermocellum celA gene encoding endoglucanase A is expressed in Saccharomyces cerevisiae but the enzyme produced from the native celA gene is not secreted. After removal of the bacterial signal peptide-coding sequence, the gene was fused to the promoter and prepro segment of the S. cerevisiae MFα1 gene. This construction directs secretion of active endoglucanase A into the culture medium when introduced in yeast on either replicating or integrating vectors. Secretion of endoglucanase A required growth of transformants on rich medium. The secreted enzyme is a 97 000 Da glycoprotein containing about half of its molecular weight as carbohydrate. This new gene fusion could facilitate further research on protein secretion in yeast by using a cellulase as a marker enzyme.
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  • 152
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    Yeast 5 (1989), S. ii 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 153
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    Yeast 5 (1989), S. 307-319 
    ISSN: 0749-503X
    Keywords: Yeast ; proton pump ; hygromycin B ; gene promoter ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Two mutations containing insertions and deletions in the promoter in the plasma membrane H+-ATPase gene (PMA1) of Saccharomyces cerevisiae have been introduced into the genome by homologous recombination, replacing the wild-type gene. The resulting strains have 15 and 23% of the wild-type ATPase content. Decreased levels of ATPase correlate with decreased rates of proton efflux and decreased uptake rates of amino acids, methylamine, hygromycin B and tetraphenylphosphonium. This supports a central role of the enzyme in yeast bioenergetics. However, the final accumulation gradient of tetraphenylphosphonium is not affected by the mutations and that of methylamine and 2-aminoisobutyric acid is only decreased in the most extreme mutant. Apparently, kinetic constraints seem to prevent the equilibration of yeast active transports with the electrochemical proton gradient. As expected from their transport defects, the ATPase-deficient mutants are more resistant to hygromycin B and more sensitive to acidification than wild-type yeast. Mutant cells are very elongated, suggesting a structural role of the ATPase in the yeast surface.
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  • 154
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    Yeast 5 (1989), S. 321-403 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 155
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    Yeast 5 (1989) 
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    Keywords: Life and Medical Sciences ; Genetics
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  • 156
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    Yeast 5 (1989), S. 429-438 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 157
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    Yeast 5 (1989), S. 439-457 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 158
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    Yeast 5 (1989), S. 405-427 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Additional Material: 6 Ill.
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  • 159
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    Yeast 5 (1989), S. 459-467 
    ISSN: 0749-503X
    Keywords: Septum ; cell wall ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Saccharomyces cerevisiae produces two chitin synthases (Chs1 and Chs2) encoded by separate genes. Although these enzymes catalyze the same reaction, Chs2 is essential for septum formation whereas Chs1 has a repair function. To determine if these physiological differences are reflected in the enzyme structures, the CHS2 gene was sequenced and compared to that of CHS1. The predicted amino acid sequence of Chs2 shares substantial similarity with that of Chs1 in the carboxyl two-thirds of the protein. The amino one-third segments differ in predicted isoelectric point by almost 5 pH units. It is suggested that the similar regions are related to common catalytic function. The unrelated regions may be involved in regulation or localization of the respective enzymes. CHS1 and CHS2 are unlinked but may have arisen from the duplication of an ancestral gene.
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  • 160
    ISSN: 0749-503X
    Keywords: Schwanniomyces castellii ; hexokinase ; continuous culture ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A mutant of Schwanniomyces castellii with reduced glucose phosphorylation and with practically no phosphorylation of fructose was investigated. Carbon catabolite represion of α-glucosidase and amylases was reduced. Repression of β-galactosidase was normal. We have compared in continuous culture this mutant strain with wild type and another previously described mutant. The relationship between the specific rate of glucose consumption (Qs) and residual glucose concentration (s) in an inverse mode, suggests that there may be two types of transport of glucose. Mutation at the phosphorylation level causes apparent modification of the kinetic parameters of glucose uptake rate. The consequence of mutation at the phosphorylation level on biomass production was discussed.
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  • 161
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    Yeast 5 (1989), S. 477-486 
    ISSN: 0749-503X
    Keywords: Schizosacchromyces pombe ; nitrogen saturation ; heat shock ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: When cells of the fission yeast, Schizosaccharomyces pombe, are incubated in medium devoid of a nitrogen source, they accelerate into cell division and differentially synthesize two polypeptides at 46 and 27 kD (named p46 and p27) after a delay of about an hour. The synthesis of p46 and p27 is transient. These proteins have no obvious cell cycle connection since they are also evident in nitrogen-starved (but not accelerated) cells of the temperature-sensitive mutant of S. pombe, wee 1·50h-. We infer from this that p46 and p27 are synthesized as a direct result of nutritional stress. The possibility that p46 and p27 represent examples of general environmental stress proteins was investigated by comparing nitrogen starvation with the heat-shock response in S. pombe. Heat-shock analysis of cells revealed the existence of two proteins of similar Mr to p46 and p27. In addition, nitrogen-starved cells acquired thermotolerance in a manner similar to heat-shocking cells. We suggest that nitrogen starvation in fission yeast induces a subset of the total array of heat-shock proteins.
    Additional Material: 8 Ill.
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  • 162
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 487-496 
    ISSN: 0749-503X
    Keywords: Flocculation ; yeast ; calcium ; cations ; efflux ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Expression of flocculation in yeast requires the presence of multi-charged cations. Calcium ions fulfil this role over a broad pH range. At near neutral pH, magnesium and a variety of transition elements can induce flocculation. Calcium-induced flocculation was competitively inhibited by excess sodium ions whereas magnesium-induced flocculation was not competitively inhibited. Potassium and lithium ions caused similar inhibition but to a lesser extent. 45Ca effux from yeast cells was greatly increased by the external presence of buffer and cations. Inhibition of flocculation by chelating agents, EDTA or EGTA, was overcome by excess calcium ions. Flocculation could not be induced, under these conditions, by excess magnesium or transition-element salts.It was concluded that yeast flocculation has a direct specific requirement for calcium ions. Other ions cause flocculation indirectly by effecting calcium ion leakage from cells, which is then able to initiate flocculation. Low calcium concentrations were susceptible to competitive inhibition by sodium ions present in most acidic buffers. In addition, citrate ions inhibited flocculation, probably by sequestration of leaked calcium. Flocculation by salts, other than calcium, was thus restricted to a neutral or high pH range.
    Additional Material: 8 Ill.
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  • 163
    ISSN: 0749-503X
    Keywords: S. cerevisiae ; Schistosoma mansoni ; immunogenic antigen ; high level of expression ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Strains of Saccharomyces cerevisiae expressing P28-I, an antigen inducing protection against schistosomiasis, have been constructed. Transformants containing a very high copy number of a P28-I expression vector were selected by genetic complementation involving deficient LEU2 or URA3 alleles carried by plasmids. Using the ura3 fur1 auto-selection system, constitutive and stable expression of P28-I could be obtained in cultures grown in rich medium.The accumulation of the foreign protein exceeds 25% of total yeast proteins when estimated by Coomassie Brilliant Blue staining of SDS-PAGE. Moreover, P28-I which was located intracellularly was soluble and biologically active.
    Additional Material: 6 Ill.
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  • 164
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989) 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 165
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    Yeast 5 (1989), S. S1 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 166
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. S213 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 167
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    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 5 (1989), S. 509-524 
    ISSN: 0749-503X
    Keywords: Cell division cycle ; DNA replication ; molecular cloning ; DNA sequence ; yeast ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A new complementation group of temperature-sensitive mutants of the yeast Saccharomyces cerevisiae (ts26-1 and ts26-2) has been isolated and characterized. This mutation maps at 40·7 cM from arg8 and 48·9 cM from arg1 on the left arm of chromosome XV of yeast, providing that it is a newly identified gene. The dumbbell-shape terminal morphology of the mutant cells at the restrictive temperatures is a characteristic of mutants defective in DNA replication. To study the defect of macromolecule synthesis in the mutant cells, DNA, RNA, and protein synthesis were measured at both permissive and restrictive temperatures. The data suggest that the primary defect of this mutation is at the initiation step of DNA synthesis.The gene has been cloned from an S. cerevisiae genomic library by rescue of the conditional lethality of the mutants. It is present as a single copy in the haploid genome. DNA-RNA hybridization of the gene has identified 1 kb RNA, which is under cell-division-cycle control. DNA sequence analysis of the gene has identified an open reading frame capable of encoding a protein of molecular weight 25 055 (214 amino acids).
    Additional Material: 9 Ill.
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  • 168
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    Yeast 5 (1989), S. S245 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 169
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    Yeast 5 (1989), S. S405 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 170
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    Yeast 5 (1989), S. S471 
    ISSN: 0749-503X
    Keywords: Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 171
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Penicillium funiculosum NRRL 13033 produced β-glucosidase and β-xylosidase activities when grown on wheat straw. The addition of some inducers (individually or in combination) to the fermentation medium were tested for the production of both enzymes. The relation of mycelial bound enzyme to cell free enzyme was studied during incubation period of fermentation. The optimum activity of β-glucosidase and β-xylosidase were found to be in the pH 4.5 using phosphate-citrate buffer at 50°C for 60 min and at 55°C for 40 min respectively. β-Glucosidase lost about 40% of its original activity by heating to 65°C for 60 min, while, β-xylosidase activity was found to be nearly stable with the same treatment. Both enzyme activities were greatly inhibited when 1.0% (w/v) of xylose and glucose were added to the assay mixture.
    Additional Material: 5 Ill.
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  • 172
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    Acta Biotechnologica 9 (1989), S. 362-362 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 173
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    Acta Biotechnologica 9 (1989), S. 361-361 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 174
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: We report here the construction of a promoter-probe vector, pRS2, which can be utilized in either Acetobacter methanolicus MB 58 or Escherichia coli due to the presence of broad-host-range replicon RSF 1010. The vector provides several unique restriction sites for promoter cloning as well as resistance markers for the selection of transformants. The promoter-probe vector was constructed by inserting an EcoRI-SalI-polylinker fragment of pUC 19 into EcoRI/SalI digested pMK 16. The resulting plasmid, pRS1, was cloned into the unique EcoRI site of the broad-host-range plasmid RSF 1010.The vector was used to clone promoter-containing sequences derived from the A. methanolicus MB 58 chromosome as well as the E. coli lac-promoter.
    Additional Material: 3 Ill.
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  • 175
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    Acta Biotechnologica 9 (1989), S. 233-238 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: A technology for the fractionation of the small-value fish species Black Sea minnow (Spratus spratus phalericus) has been proposed. Four valuable fractions with a possibility for utilisation as food and health products have been proposed: the protein isolate, the aminoacid concentrate, the lipid fraction and the fish-bone fraction.
    Additional Material: 1 Ill.
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  • 176
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    Acta Biotechnologica 9 (1989), S. 247-253 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The sorption capacity of silver on different biological materials has been investigated depending on physico-chemical pretreatments. The maximum silver loading values measured were compared with the values obtained with nontreated biomasses.The results show an increase of the loading capacity up to a factor of 10 in case of the alkalitreated biomasses. When the biomasses are extracted before being used as adsorbent with a solvent mixture of chloroform/methanol in a ratio of 2:1 the efficiency of the silver adsorbing power can be increased.Beyond that, the ability to adsorb silver can also be influenced when microorganisms are used as biocatalysts in a product synthesis before they are used as adsorbents. A strain of Acetobacter methanolicus possesses 1.8 times higher affinity to silver when it is employed in a process of gluconic acid production before adsorption. Physico-chemical pretreatments influence not only the loading capacity of the biological material, but also the contacting time required for the establishment of the adsorption equilibrium can be considerable reduced.
    Additional Material: 4 Ill.
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  • 177
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    Acta Biotechnologica 9 (1989), S. 268-268 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 178
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    Acta Biotechnologica 9 (1989), S. 383-387 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: UV/VIS diffuse reflectance spectroscopy and fluorescence spectroscopy have been used to investigate the cytochrome and pyridine nucleotide spectra during aerobic biomass growth of Saccharomyces cerevisiae followed by an anaerobic ethanol formation process. The cytochrome and NAD(P)H spectra are closely related to fermentation parameters such as biomass growth rate and ethanol concentration.
    Additional Material: 4 Ill.
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  • 179
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 180
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The selective and complementary interaction between the ligand bound to support surface and isolated substance is the key of the affinity chromatography. This is the reason of the research in the field of certain sorbents. They are obtained by chemical reaction of matrix with proper ligands. It is carried out mainly by using bifunctional substances which make the reaction with the ligand in question possible. This paper deals with the preparation of controlled porosity glass with reactive epoxy groups. In order to obtain such support and avoid application of a proper epoxy silane compound, the double-step reaction using simple agents was employed. The sorbent syntheses were optimized. The prepared sorbents were applied for coupling some carbohydrates and amino acids.
    Additional Material: 3 Ill.
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  • 181
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    Acta Biotechnologica 9 (1989), S. 294-294 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 182
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    Acta Biotechnologica 9 (1989), S. 295-298 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The aim of this work is to study the IR spectra of the different lignins precipitated from the produced liquor of different pulping conditions of bagasse and cotton stalks, also to study their antimicrobial activities towards some bacteria and fungi.
    Additional Material: 1 Tab.
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  • 183
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    Acta Biotechnologica 9 (1989), S. 291-293 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: A mutant of Corynebacterium glutamicum dependent on homoserine and resistant both to S-(2-aminoethyl)-L-cysteine and lysine hydroxamate, cultivated under submerged conditions for 4 days in a medium containing sucrose, corn-steep and pea nut meał hydrolyzate, accumulated 33.5 g/l lysine.
    Additional Material: 2 Tab.
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  • 184
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    Acta Biotechnologica 9 (1989), S. 324-324 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 185
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    Acta Biotechnologica 9 (1989), S. 346-346 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 186
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    Acta Biotechnologica 9 (1989), S. 325-332 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: In the present work, a mathematical model was developed regarding the immobilized living yeast cell reactor for sugar bioconversion to ethanol. The model, composed of a system of ordinary differential equations (ODEs) enables the computation of the paramters involved in the steady state reactor behaviour. Comparing the values computed through the integration of this mathematical model with the experimental data, it has been shown its capacity to describe sufficiently accurate the steady state behaviour of the continuous fixed film bioconversion reactor.
    Additional Material: 7 Ill.
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  • 187
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    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 188
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 189
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 190
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The increased interest in large scale production of biologically active molecules, as for example monoclonal antibodys, hormones, proteins and enzymes, has stimulated a rapid development of different methods to cultivate eukaryotic cells. Further progress in this field of modern biotechnology is expected not only from the selection of more productive cell lines and more efficient cultivation techniques, but also from the improvement in new bioreactor design and operation, which guarantees increased productivity per unit volume and reduces the downstream processing. Most important factors for a new reactor design in future will be the energy- and mass transfer, shear stress and scaleability. The provision of an adequate oxygen supply to large scale reactors is the most critical barrier to scale up. If oxygen is limited in a small degree, the result is inhibition of cell density and cellular efficiency in the production of the desired biomolecules. In addition, when methodologies are used which allow high cell densities and high metabolic active cells, the oxygen transfer becomes more important and, at the same time, more difficult. Since direct sparging of air into the cell-containing medium causes problems such as shear forces and foaming, new, efficient methods in bubble free aeration must be utilized. A new aeration system is presented, which is suitable to bubble-free aeration as well to separation of microcarrier-anchered cells from the harvested medium when running in a continuous perfusion mode. The efficiency of the CHEMCELL System regarding aeration and cell retention is demonstrated by the growth kinetics of BHK 21-cells in batch and perfusion mode.
    Additional Material: 6 Ill.
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  • 191
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The MODELBASE BIOTECHNOLOGY has been developed for gathering and processing data obtained from biochemical experiments. It covers the analysis of growth kinetics and production kinetics as well and may be used for calibration curve determination too. The implementation of models defined by the user is supported. Both, differential equations and their integrated forms are accepted. In addition to parameter estimation a statistical analysis can be carried on request. Two and three dimensional graphical representation facilitates the interpretation of results.Fields of application are biochemistry, biotechnology, microbiology, engineering, pharmacy, equipment design, education.
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  • 192
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    Acta Biotechnologica 9 (1989), S. 3-8 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Although the regulation of microbial secondary metabolism belongs to the important objects of actual investigations the results and knowledges are used rather poorly in industrial fermentations. This situation arises from the discrepancy between the economically driven development of the know how as soon as possible contrary to the more slow progress of profound examination of the complicated network of secondary metabolism. Nevertheless some well known principles of metabolic regulation, e.g. catabolite repression or resistance against products or metabolites are considered in fermentation processes by means of suitable substrate application as well as improvement of high yield strains. Both aspects are discussed with respect to examples of fermentations of β-lactams, polyketides and glycopeptides.
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  • 193
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: The loss of fermentative activity of yeast cells, observed in continuous fermentation experiments at increasing biomass concentration is explained by the assumption that the ethanol-tolerance behaviour of the microorganisms changes if a growth-stabilizing factor limitation is present. A mathematical specification of the relationships existing in this context is given and an improved steady-state productivity model of ethanol production is derived.
    Additional Material: 3 Ill.
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  • 194
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 195
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: A controlled brew of pito was produced and compared with a native brew. The former contained a higher alcohol content than the latter brew and was brighter in colour (13 as against 18.5 EBC units) thus making it more attractive. Of various heat and chemical preservative treatments used to prolong the shelf life of both products, pasteurization at 75°C for 30 min. and treatment with sorbic acid at a concentration of 5% were best able to arrest microbial proliferation in the bottled products during a 4 week storage period.
    Additional Material: 2 Tab.
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  • 196
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    Acta Biotechnologica 9 (1989), S. 143-148 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Nine Candida species isolated from fleshy fruits have been examined for their organic growth factor requirements. Complete or partial deficiencies for thiamine, biotin, pyridoxine, inositol, pantothenic acid, ascorbic acid and xanthine have been reported in C. catenuclta, C. curiosa, C. curvata, C. humicola, C. rhagii, C. steatolytica and C. tropicalis. Only two species, namely, C. fragicola and C. ishiwadea were prototrophic for the growth factors. Concentrations of essential growth factors higher than the optimum have been found inhibitory for the growth of yeasts.
    Additional Material: 1 Ill.
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  • 197
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    Acta Biotechnologica 9 (1989), S. 177-177 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 198
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 199
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    Acta Biotechnologica 9 (1989) 
    ISSN: 0138-4988
    Keywords: Life Sciences ; Life Sciences (general)
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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  • 200
    ISSN: 0138-4988
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    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
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