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  • 1
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Multidrug resistance (MDR) in bacteria has been associated with efflux pumps that export structurally unrelated compounds and decrease cytoplasmic drug accumulation. To investigate MDR in mycobacteria, we studied the Mycobacterium smegmatis mutant mc211, which is resistant to doxorubicin, tetracycline, rhodamine, ethidium bromide and the hydrophilic fluoroquinolones. A genomic library constructed from this mutant was used to select clones conferring resistance to doxorubicin. Surprisingly, the clone selected encodes the efflux pump LfrA, which has been reported to confer resistance to hydrophilic fluoroquinolones, ethidium bromide, rhodamine, and acriflavine. To define the contribution of LfrA to the innate mycobacterial drug resistance and to the MDR phenotype in mc211, the lfrA gene was disrupted in both the mc211 mutant and the mc2155 wild-type parent. LfrA disruption of the wild-type strain decreased resistance to ethidium bromide and acriflavine, and increased accumulation of ethidium bromide. However, disruption of lfrA gene results only in a 2-fold decrease in minimal inhibitory concentrations (MICs) for ciprofloxacin, doxorubicin, rhodamine, and accumulation of [14C]ciprofloxacin was unchanged. LfrA disruption of the MDR strain mc211 produced a similar phenotype. Thus, LfrA contributes significantly to the intrinsic MICs of M. smegmatis for ethidium bromide and acriflavine, but not for ciprofloxacin, doxorubicin or rhodamine.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The genus Mycobacterium is composed of species with widely differing growth rates ranging from approximately three hours in Mycobacterium smegmatis to two weeks in Mycobacterium leprae. As DNA replication is coupled to cell duplication, it may be regulated by common mechanisms. The chromosomal regions surrounding the origins of DNA replication from M. smegmatis, M. tuberculosis, and M. leprae have been sequenced, and show very few differences. The gene order, rnpA-rpmH-dnaA-dnaN-recF-orf-gyrB-gyrA, is the same as in other Gram-positive organisms. Although the general organization in M. smegmatis is very similar to that of Streptomyces spp., a closely related genus, M. tuberculosis and M. leprae differ as they lack an open reading frame, between dnaN and recF, which is similar to the gnd gene of Escherichia coli. Within the three mycobacterial species, there is extensive sequence conservation in the intergenic regions flanking dnaA, but more variation from the consensus DnaA box sequence was seen than in other bacteria. By means of subcloning experiments, the putative chromosomal origin of replication of M. smegmatis, containing the dnaA-dnaN region, was shown to promote autonomous replication in M. smegmatis, unlike the corresponding regions from M. tuberculosis or M. leprae.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 49 (1988), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Growth of the cyanobacterium Spirulina platensis, like that of many prokaryotic and eukaryotic organisms, is inhibited by low concentrations of valine, one of the three end-products of the branched-chain amino acid biosynthetic pathway. We assayed and partially characterized the activity of acetyhydroxy acid synthase (AHAS), the first common enzyme of the branched pathway in cell-free extracts from axenic S. platensis cultures. Assays performed at various pH values showed two peaks of activity, both inhibited by valine. FAD was not required for enzyme activity but protected it during dialysis. We also investigated whether the three amino acids were able to cause repression of AHAS synthesis and a significant drop in the enzyme-specific activity could be seen only when cultures were grown in the presence of valine. Chromatography on hydroxylapatite showed one single peak of activity.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 49 (1988), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Growth of the cyanobacterium Spirulina platensis is inhibited by low concentrations of valine. Spontaneous valine-resistant mutants were isolated from S. platensis and a preliminary characterization of three of them indicates that one (strains DR2) is defective in valine uptake and two (strains DR5 and DR9) carry alterations in a valine-mediated mechanism of synthesis of acetohydroxy acid synthase, the first common enzyme of the pathway.
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 30 (1985), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract A Spirulina platensis gene library has been constructed using cosmid vector pMMB34. The cosmid bank was controlled for its random gene distribution by colony hybridization. Genes were identified using either homologous or heterologous probes of genes involved in photosynthesis (large and small subunit of d-ribulose 1,5-bisphosphate carboxylase, 32 kDa thylakoid protein, α, β subunits of C-phycocyanin) and protein synthesis (elongation factors EF-Tu, EF-G).
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-072X
    Keywords: Acetohydroxy acid synthase ; Acetolactate synthase ; Isozymes ; Isoleucine-leucine-valine biosynthesis ; Spirulina ; Cyanobacteria ; Flavin adenine dinucleotide requirement
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Two isoforms of acetohydroxy acid synthase (AHAS), the first enzyme of the branched-chain amino acids biosynthetic pathway, were detected in cell-free extracts of the cyanobacterium Spirulina platensis and separated both by ion-exchange chromatography and by hydrophobic interaction. Several biochemical properties of the two putative isozymes were analysed and it was found that they differ for pH optimum, FAD requirement for both activity and stability, and for heat lability. The results were partially confirmed with the characterization of the enzyme extracted from a recombinant Escherichia coli strain transformed with one subcloned S. platensis coli strain transformed with one subcloned S. platensis AHAS gene. The approximate molecular mass of both AHAS activities, estimated by gel filtration, indicates that they are distinct isozymes and not different oligomeric species or aggregates of identical subunits.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-072X
    Keywords: Acetohydroxy acid synthase ; Ilv genes ; Cyanobacteria ; Isoleucine ; Leucine ; Valine biosynthesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The coding sequence for Spirulina platensis acetohydroxy acid synthase (AHAS, EC 4.1.3.18) is shown to be contained within a 4.2 Kb ClaI fragment (ilvX) that has been cloned from a recombinant lambda library. This fragment was able to complement a suitable mutant of Escherichia coli when inserted into the ClaI site of plasmid pAT153 in either orientation, demonstrating that transcription of ilvX originated within the cloned fragment. The probe used for hybridization experiments was the corresponding gene from Anabaena sp. PCC7120. The same probe allowed us to identify a second putative gene encoding AHAS in the S. platensis genomic library.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Plant molecular biology 4 (1985), S. 133-136 
    ISSN: 1573-5028
    Keywords: cloning ; glutamine synthetase gene ; Spirulina platensis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract An 8 Kilobase-pair (Kbp) HindIII fragment containing the coding sequence forSpirulina platensis glutamine synthetase [EC 6.3.1.1.] has been identified utilizing a probe derived fromAnabaena 7120 and cloned in the vector pAT153.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Screening for the plasmid content of 94 strains belonging to ten species of thermophilic bacilli mainly referred toBacillus stearothermophilus was carried out. Twenty-seven of the strains tested were found to harbor one or more plasmids ranging in size from 1.7 to〉50 kb. The physical map of the smallest plasmid, pBC1, is reported. Southern hybridization experiments showed that pBC1 hybridized strongly with the other small plasmids, weakly with medium-sized plasmids, and not at all with the largest plasmids and chromosomal DNA. pBC1 was cloned into pHV14 and pJH101 vectors, and the chimeric plasmids obtained were used to transformEscherichia coli andBacillus subtilis.
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1573-6784
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Summary Transformation of intact cells of Bacillus licheniformis 5A24 with plasmids pLM6 (2.8 kb), pC194 (2.9 kb) and pCP49 (7.1 kb) by electroporation resulted in 1.5 × 106, 1.2 × 106 , and 5.2 × 104 transformants per μg DNA, respectively. Transformation was possible with plasmid DNA, which was religated after restriction endonuclease digestions. In addition, evidence is presented that indicates that B. licheniformis possesses DNA restriction and modification systems.
    Type of Medium: Electronic Resource
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