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  • 1
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Plasmid pCIT264 from Lactococcus lactis subsp. lactis biovar diacetylactis (L. diacetylactis) contains an insertion sequence (IS)-like element located in the citrate utilization (citQRP) cluster. This 967-nucleotide long element is bounded by 17 bp perfect inverted repeats and contains an open reading frame (ORF1) composed of 296 codons, which could encode a transposase. Expression of the IS from pCIT264 generates two mRNAs of 2900 and 1900 nucleotides. The transcription is driven by the P3 promoter, composed of a — 10 region located at the right end of the IS and of a — 35 region positioned downstream of this element. The IS-like element (IS982) is present in seven copies in the L. diacetylactis genome. The copy present in pCIT264 is highly stable and does not promote rearrangements of the cit cluster. We suggest that the stable maintenance of the IS-like element in pCIT264 could be due to a translational control of the putative transposase by an antisense RNA.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract The Lactococcus lactis subsp. lactis biovar diacetylactis citrate permease P is encoded by the citP gene borne by an 8.3-kb Cit+ plasmid (F. Sesma, Gardiol, D., de Ruiz Holgado, A.P. and de Mendoza, D., Appl. Environ. Microbiol. 56 (1990) 2099–2103). By subcloning different DNA fragments from the Cit+ plasmid it was found that citP is the only plasmidic gene required for the transport of citrate. Northern blot analysis revealed that citP is transcribed in two mRNA species with lengths of 2.9 and 1.9 kb. The transcriptional pattern, the total amount of citP mRNAs and the transport of citrate were unaffected by the presence of citrate in the growth media. Our results support the conclusion that transcription of citP as well as citrate transport are not induced by citrate in Lactococcus lactis subsp. lactis biovar diacetylactis.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 11 (1994), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Bacillus subtilis growing at 37° C synthesizes, almost exclusively, saturated fatty acids. However, when a culture growing at 37°C is transferred to 20°C, the synthesis of unsaturated fatty acids is induced. The addition of the DNA gyrase inhibitor novobiocin specifically prevented the induction of unsaturated fatty acid synthesis at 20° C. Furthermore, it was determined that plasmid DNA isolated from cells growing at 20°C was significantly more negatively supercoiled than the equivalent DNA isolated from cells growing at 37°C. The overall results agree with the hypothesis that an increase in DNA supercoiling associated with a temperature downshift could regulate the unsaturated fatty acids synthesis in B. subtilis.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 8 (1993), S. 0 
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Bacillus subtilis synthesizes, almost exclusively, saturated fatty acids, when grown at 37° C. When cultures were transferred from 37° C to 20° C, a chloramphenicol- and rifampicin-sensitive synthesis of a C-16 unsaturated fatty acid was observed. Synthesis of this compound reached a plateau after 5 h at 20° C, reaching levels of 20% of the total fatty acid content. [14C]-labelled fatty acids attached as thioesters to acyl-carriers compounds, such as coenzyme A (CoA) or acyl-carrier protein (ACP) synthesized de novo by glycerol-requiring auxotrophs deprived of glycerol to arrest phospholipid synthesis, could not be desaturated at 20° C. Desaturation of these fatty acids was readily observed when glycerol was restored to the cultures allowing resumption of transfer of acyl-moieties from acyl-thioesters to phospholipid. It was also observed that depletion of the pools of CoA and ACP by starvation of pantothenate auxotrophs had no effect on the observed synthesis of unsaturated fatty acid at 20° C. The overall results indicate that synthesis of unsaturated fatty acids in B. subtilis is a cold-inducible process and that phospholipids are obligate intermediates in this fatty acid desaturation pathway.
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  • 5
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The citMCDEFGRP cluster from Leuconostoc paramesenteroides involved in citrate utilization was cloned and its nucleotide sequence determined. Homology of the inferred gene products with characterized enzymes reveals that citP encodes the citrate permease P, citC the citrate ligase and citDEF the subunits of the citrate lyase of Leuconostoc. Moreover, it suggests that citM encodes a Leuconostoc malic enzyme. Analysis of citrate consumption by and citrate lyase activity of Lc. paramesenteroides J1[pCITJ1] showed that its citrate permease and its citrate lyase are induced by the presence of citrate in the growth medium. Southern blot analysis demonstrated that the citMCDEFGRP cluster is located in a plasmid.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 142 (1996), S. 0 
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract Citrate uptake in Lactococcus lactis ssp. lactis biovar diacetylactis (L. diacetylactis) is catalysed by a secondary citrate carrier, the citrate permease P (CitP). In this report the mechanism of citrate uptake was investigated in resting cells of L. diacetylactis. The results presented reveal that the pH gradient (ΔpH) and the membrane potential (ΔΨ) are driving forces for citrate uptake. Moreover, it seems that CitP activity requires neither Na+ nor Mg2+ cations.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Summary The Des pathway of Bacillus subtilis regulates the expression of the acyl-lipid desaturase, Des, thereby controlling the synthesis of unsaturated fatty acids (UFAs) from saturated phospholipid precursors. Previously, we showed that the master switch for the Des pathway is a two-component regulatory system composed of a membrane-associated kinase, DesK, and a soluble transcriptional regulator, DesR, which stringently controls transcription of the des gene. Activation of this pathway takes place when cells are shifted to low growth temperature. Here, we report on the mechanism by which isoleucine regulates the Des pathway. We found that exogenous isoleucine sources, as well as its α-keto acid derivative, which is a branched-chain fatty acid precursor, negatively regulate the expression of the des gene at 37°C. The DesK–DesR two-component system mediates this response, as both partners are required to sense and transduce the isoleucine signal at 37°C. Fatty acid profiles strongly indicate that isoleucine affects the signalling state of the DesK sensor protein by dramatically increasing the incorporation of the lower-melting-point anteiso-branched-chain fatty acids into membrane phospholipids. We propose that both a decrease in membrane fluidity at constant temperature and a temperature downshift induce des by the same mechanism. Thus, the Des pathway would provide a novel mechanism to optimize membrane lipid fluidity at a constant temperature.
    Type of Medium: Electronic Resource
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  • 8
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The initiation of sporutation in Bacillus subtilis is controlled by the Spo0A transcription factor which is activated by phosphorylation through a phosphorelay mechanism that is dependent upon the activity of one or more protein kinases. The enzymatic activity of one of these protein kinases, KinA, was found to be inhibited in vitro by certain fatty acids. The most potent inhibitors have at least one unsaturated double bond in the cis configuration and a chain length of 16–20 carbon atoms. Homologous isomers with a trans double bond are not inhibitory. Saturated straight- or branched-chain fatty acids are either much weaker inhibitors or have no effect. The inhibitors prevent autophosphorylation of KinA and are non-competitive with ATP. B. subtilis phospholipids were found to contain at least one as yet unidentified type of fatty acid that, when present in an unesterified form, inhibited KinA. The results suggest that the concentration of a specific unsaturated fatty acid may act as a signal linking the initiation of sporulation to the status of membrane synthesis and septation or some other specific membrane-associated activity.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: A hallmark of sporulation of Bacillus subtilis is the formation of two distinct cells by an asymmetric septum. The developmental programme of these two cells involves the compartmentalized activities of σE in the larger mother cell and of σF in the smaller prespore. A potential role of de novo lipid synthesis on development was investigated by treating B. subtilis cells with cerulenin, a specific inhibitor of fatty acid biosynthesis. These experiments demonstrated that spore formation requires de novo fatty acid synthesis at the onset of sporulation. The transcription of the sporulation genes that are induced before the formation of two cell types or that are under the exclusive control of σF occurred in the absence of fatty acid synthesis, as monitored by spo–lacZ fusions. However, expression of lacZ fusions to genes that required activation of σE for transcription was inhibited in the absence of fatty acid synthesis. The block in σE-directed gene expression in cerulenin-treated cells was caused by an inability to process pro-σE to its active form. Electron microscopy revealed that these fatty acid-starved cells initiate abnormal polar septation, suggesting that de novo fatty acid synthesis may be essential to couple the activation of the mother cell transcription factors with the formation of the differentiating cells.
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  • 10
    ISSN: 1617-4623
    Keywords: Key words Transcriptional activation ; Lactococcuslactis ; Escherichia coli ; Plasmid pCIT264 ; Expression of citP gene
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract  The lactococcal plasmid pCIT264 contains a cluster of three genes (citQ, citR and citP) involved in the transport of citrate in Lactococcus lactis biovar diacetylactis. The cit cluster contains a copy of a newly discovered insertion sequence (IS)-like element located between its promoter P1 and the first gene of the cluster. In this report, we show that this IS-like element can act as a mobile switch for the downstream genes, creating two new transcriptional promoters named P2 and P2′. The P2 promoter is recognized by the lactococcal RNA polymerase in vivo. This is a hybrid promoter composed of a −35 region reading outwards 12 bp from the right end of the IS-like element, and a nucleotide sequence from the recipient plasmid, adjacent to the element, which provides an appropriately spaced −10 region. Transcription of the citQRP cluster from this promoter takes place during the exponential and stationary phases of growth in L. lactis. Promoter P2′ is included in the IS-like element and is the only promoter responsible for expression of citP in E. coli. Thus, it appears that the introduction of this element into pCIT264 allows expression of the citQRP cluster in E. coli, and increases its levels of expression in L. lactis.
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