bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequence analysis of the wall‐associated protein precursor of Streptococcus mutans antigen A
Authors: Ferretti J. J. , Russell R. R. B. , Dao M. L. 
Published: October 2006

SummaryThe nucleotide sequence has been determined for the Streptococcus mutans wall‐associated protein A (wapA) gene from serotype c strains Ingbritt and GS5. The nucleotide sequence for each wapA gene was virtually identical, although the gene from strain GS5 contained a 24 base pair deletion. A 29 amino acid signal peptide was specified by each wapA gene with a mature protein of 424 amino acids (Mr, 45276) for strain Ingbritt and 416 amino acids (Mr, 44846) for strain GS5. In the C‐terminal region of the wall‐associated protein A, considerable sequence similarity was found with the membrane anchor region of proteins from other Gram‐positive organisms such as the group A streptococcal M protein and the group G streptococcal IgG binding protein. Adjacent to the proposed membrane anchor is a highly hydrophilic region which may span the cell wall; both sequence data and experimental evidence indicate the existence of a region immediately outside the wall at which proteolytic cleavage occurs to release antigen A of Mr 29000 into the culture supernatant. Thus, the wall‐associated protein A is a precursor of the 29000 Mr antigen A.
Klebsiella pneumoniae strain K21: evidence for the rapid secretion of an unacylated form of pullulanase
Authors: Kornacker M. G. , Boyd A. , Pugsley A. P. , Plastow G. S. 
Published: October 2006

Summary Klebsiella pneumoniae strain PAP996 was previously shown to secrete fatty acylated, aggregated (micellar) pullulanase only after the end of exponential growth. Here we show that the closely related strain K21 secretes large amounts of unacylated, non‐aggregated (monomeric) pullulanase during exponential growth. Only a small amount (<10%) of the secreted pullulanase was initially retained by the exponentially growing cells to be subsequently secreted in a fatty acylated, aggregated form. Despite the absence of fatty acids in secreted monomeric pullulanase, the effects of the antibiotic globomycin on pullulanase maturation indicated that all of the enzyme synthesized by strain K21 is processed by lipoprotein signal peptidase.
Characterization of the neisserial lipid‐modified azurin bearing the H.8 epitope
Authors: Woods J. P. , Dempsey J. F. , Kawula T. H. , Barritt D. S. , Cannon J. G. 
Published: October 2006

SummaryThe pathogenic Neisseria have multiple genes encoding proteins that bind monoclonal antibody (MAb) H.8. We previously reported the cloning and sequencing of a meningococcal gene (laz) encoding an H.8 Mab‐binding protein with a consensus lipoprotein processing site, an N‐terminal domain containing the epitope for H.8 MAb binding, and a C‐terminal domain with extensive similarity to the sequences of azurins from other organisms. In the current study, we showed that the product of the cloned gene could be labelled with palmitic acid, that it was subject to globomycin‐sensitive processing, and that it was immunologically cross‐reactive with azurin from Pseudomonas aeruginosa. All neisserial species tested, both pathogens and commensals, produced a protein recognized by anti‐azurin serum. Southern blots with oligonucleotide probes specific for the azurin domain of the gene showed that it was present in a single copy in the chromosome; it was highly conserved in gonococci and meningococci, and less conserved in commensal Neisseria species.
Construction of lac fusions to the inducible arginine‐and lysine decarboxylase genes of Escherichia coli K12
Authors: Auger E. A. , Redding K. E. , Plumb T. , Childs L. C. , Meng S.‐Y. , Bennett G. N. 
Published: October 2006

SummaryThe induction of several amino acid decarboxylases under anaerobic conditions at low pH has been known for many years, but the mechanism associated with this type of regulation has not been elucidated. To study the regulation of the biodegradative arginine and lysine decarboxylases of Escherichia coli K12, Mudlac fusions to these genes were isolated. Mudlac fusion strains deficient for lysine decarboxylase or arginine decarboxylase were identified using decarboxylase indicator media and analysed for their regulation of β‐galactosidase expression. The position of the Mud‐lac fusion in lysine decarboxylase‐deficient strains has been mapped to the cadA gene at 93.7 minutes, while the Mudlac fusions exhibiting a deficiency in the inducible arginine decarboxylase have been mapped to 93.4 minutes.
Molecular characterization and sequence of phosphatidylinositol‐specific phospholipase C of Bacillus thuringiensis
Authors: Lechner M. , Kupke T. , Stefanovic S. , Götz F. 
Published: October 2006

SummaryThe gene encoding monophosphatidylinositol inositol phosphohydrolase (PI‐specific phospholipase C, PI‐PLC) of Bacillus thuringiensis was cloned in Staphylococcus carnosus TM300. The complete coding region comprises 987 base pairs corresponding to a precursor protein of 329 amino acids (molecular weight, 38095). The NH2‐terminal sequence of the purified enzyme from Escherichia coli indicated that the mature PI‐PLC consists of 299 amino acid residues with a molecular weight of 34586. Polyacrylamide gel electrophoresis revealed the same molecular weight for the purified enzyme isolated from the DNA‐donor strain of B. thuringiensis and from the E. coli clone. By computer analysis, the secondary structure was predicted. The enzyme from the E. coli recombinant shows no activity on other phospholipids and sphingo‐myelin. The cleaving specifity of PI‐PLC was examined by thin layer chromatography.
Topographical alterations in proteins I of Neisseria gonorrhoeae correlated with lipooligosaccharide variation
Authors: Judd R. C. , Shafer W. M. 
Published: October 2006

SummaryFour transformant strains of Neisseria gonorrhoeae were generated, two of which (WS3 and WS5) had protein I subclass A (P.IA) and two which (WS2 and WS4) had protein I subclass B (P.IB). Analysis of the strains demonstrated that the two P.IA‐bearing strains differed in lipooligosaccharide (LOS) and H.8 antigen, as assessed by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and immuno‐blotting. The WS5 strain had slow‐migrating LOS and H.8 antigen, and the WS3 strain had fast‐migrating LOS and H.8 antigen. The P.IB‐bearing strains also had either slow‐migrating LOS and H.8 antigen (WS4) or fast‐migrating LOS and H.8 antigen (WS2). Structural and exposure analysis revealed that although the P.IAs were identical in the WS3 and WS5 strains, there was a slight alteration of the exposure of the proteins which correlated with altered LOS and/or H.8 antigen. The P.IBs were also shown to be structurally identical, but the LOS and/or H.8 antigen variation in these strains correlated with a more pronounced alteration in the exposure of the P.IB molecules. The differences in protein I (P.I) exposure were generally found in highly negatively charged regions of the molecule, suggesting that the immunogenicity and/or antigenicity of the P.I molecules may vary as a result of LOS and/or H.8 antigen alterations.
The proline transport protein of Aspergillus nidulans is very similar to amino acid transporters of Saccharomyces cerevisiae
Authors: Sophianopoulou V. , Scazzocchio C. 
Published: October 2006

SummaryIn Aspergillus nidulans, the gene prnB encoding the major proline transport system is one of a cluster of four genes necessary and sufficient for the utilization of proline as sole nitrogen and/or carbon source. The prn cluster has been cloned and the sequence and transcript map of the prnB gene are presented in this paper. The predicted translated sequence consists of 570 amino acids, resulting in a molecular weight of 63028 Daltons. Its hydropathy profile shows 10 hydrophobic segments typical of integral membrane proteins. No N‐terminal hydrophobic signal peptide is present, the N‐terminal and C‐terminal ends of the protein being hydrophilic. Similar results were previously found for the arginine and histidine transporters of Saccharomyces cerevisiae, with which the prnB transporter shares regions of highly conserved amino acid sequences. Using S1 mapping and Northern blot analyses, we confirm the presence of a unique inducible prnB transcript of 1.9 kb.
A Yersinia pestis‐specific DNA fragment encodes temperature‐dependent coagulase and fibrinolysin associated phenotypes
Authors: McDonough K. A. , Falkow S. 
Published: October 2006

SummaryThe effect of temperature on coagulase and fibrinolysin expression (Pla) by Yersinia pestis has been implicated in the transmission of plague by fleas. In an attempt to improve our understanding of this process, we have cloned, sequenced and characterized the gene encoding the Pla phenotypes in Y. pestis, and examined its temperature‐dependent regulation. The coding region for this gene overlaps a 900bp Y. pestis‐specific DNA fragment that we have previously shown to be capable of detecting plague bacilli in fleas. The pla gene contains a single open reading frame encoding 312 amino acids with a predicted molecular weight of 34.7 kD and a putative signal sequence of 20 amino acids. This coding region appears to be sufficient for both coagulase and fibrinolytic activities. In Y. pestis, modulation between coagulase and fibrinolytic activities is temperature‐dependent: coagulase activity is most evident at temperatures below 30°C but fibrinolytic activity increases with higher temperatures (>30°C), regardless of the temperature at which the bacteria are grown. Our results lead us to believe that this regulation occurs post‐translationally. It is possible that the alternative forms of the Pla protein are essential to ‘flea blockage’ and subsequent transmission of the plague bacillus to animals.
Immunity protein is not required for the bactericidal activity of colicin E3
Authors: Escuyer V. , Perrin D. , Mock M. 
Published: October 2006

SummaryPlasmid pLAX3, carrying the colicin E3 gene, was used to direct the in vitro synthesis of a colicin E3*molecule totally devoid of its immunity protein. We established that this molecule is able to kill sensitive Escherichia coli cells in the total absence of immunity protein. Therefore, all of the information required for colicin E3 action is located on the colicin potypeptide itself. Furthermore, our studies indicated that immunity protein protects the C‐terminal enzymatic part of native colicin E3 protein against proteolytic degradation before or during its translocation across the cell envelope. These results are discussed in relation to the mode of entry of colicin E3 into bacterial cells.
Analysis of C4‐dicarboxylate transport genes in Rhizobium meliloti
Authors: Yarosh O. K. , Charles T. C. , Finan T. M. 
Published: October 2006

SummaryA 5.1 kbp DNA fragment was isolated which complemented C4‐dicarboxylate transport mutants (dct) of Rhizobium meliloti. Characterization of this fragment by subcloning, transposon mutagenesis, and complementation analysis revealed three loci, designated dctA, dctB, and dctD. TnphoA‐generated alkaline phosphatase fusions to dctA suggested that this gene encodes the structural transport protein and allowed the determination of its direction of transcription. Analysis of the fusions in various mutant backgrounds demonstrated that dctB, dctD, and ntrA products are required for dctA expression. The dctA fusion was constitutively expressed in a dctA mutant background, but was not expressed in dctA dctB or dctA dctD double mutants. This suggests that the constitutive expression in a dctA mutant background is mediated through dctB and dctD. Three independent second‐site Dct+ revertant mutations in ntrA mutant strains mapped to the dct locus. Succinate transport in these revertant strains was constitutive, whereas in the wild type, succinate transport was inducible. These results are consistent with the direct requirement of the ntrA gene product for dctA expression. Alfalfa plants inoculated with the dctB and dctD mutants showed reduced nitrogen‐fixing activity. Nodules induced by dctA mutants failed to fix nitrogen. These symbiotic phenotypes are consistent with previous suggestions that dctA expression in bacteroids can occur independently of dctB and dctD.
Detection and identification of mycobacteria by amplification of mycobacterial DNA
Authors: Hance A. J. , Grandchamp B. , Lévy‐Frébault V. , Lecossier D. , Rauzier J. , Bocart D. , Gicquel B. 
Published: October 2006

SummaryA 383bp segment of the gene coding for the 65kD mycobacterial antigens from Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium avium, Mycobacterium paratuberculosis, and Mycobacterium fortuitum was amplified using Tag polymerase and synthetic oligonucleotide primers and the amplified DNAs from four of these species were compared by nucleotide sequencing. Although the gene segments from these species showed considerable similarity, oligonucleotide probes which could distinguish M. tuberculosis/M. bovis, M. avium/M. paratuberculosis and M. fortuitum could be identified. Samples containing 106 human cells and serial dilutions of a suspension of intact mycobacteria were prepared, DNA was extracted, the segment of the mycobacterial DNA sequence amplified, and the amplified DNA hybridized with oligonucleotide probes. In two independent experiments, this procedure permitted the detection and identification of less than 100 mycobacteria in the original sample. These results suggest that this approach may prove useful in the early diagnosis of mycobacterial infection.
Independent and coupled translational initiation of atp genes in Escherichia coli: experiments using chromosomal and plasmid‐borne lacZ fusions
Authors: Gerstel B. , McCarthy J. E. G. 
Published: October 2006

SummaryThe translational initiation rates directed by the translational initiation regions (TIRs) of the atpB, atpH, atpA and atpG genes of Escherichia coli were investigated using lacZ fusions present on plasmids as well as integrated into the chromosome. This was the first investigation of the translational efficiency of the atpB gene, whose unfused product (subunit a) can be toxic to the cell. The specific mRNA levels, rates of in vivo protein synthesis and β‐galactosidase activities encoded by the atp:: lacZ fusions were compared in order to obtain valid estimates of relative translation rates. The results indicate that in the E. coli atp operon, translation directed by the atpB, atpH and atpG TIRs is less efficient than that directed by the atpA TIR, and are thus consistent with earlier measurements of direct atp gene expression. Initiation is, however, to differing extents, controlled by coupling to the translation of upstream neighbours. There is particularly tight coupling between atpH and atpA. Increasing the distance between these two genes whilst maintaining the original atpA TIR structure decreased the degree of coupling. The influence of manipulations of the atpG TIR structure upon translational efficiency was quantitatively more pronounced when the atpG fusions were present as a single copy per chromosome. This is likely to be related to the mRNA binding characteristics of 30S ribosomal subunits and/or to the influence of other (trans‐acting) factors. The control of independent and coupled initiation at the atp TIRs is discussed in relation to mRNA structure and possible CiS and trans regulatory phenomena.
Pseudomonas aeruginosa cytotoxin: the nucleotide sequence of the gene and the mechanism of activation of the protoxin
Authors: Hayashi T. , Kamio Y. , Hishinuma F. , Usami Y. , Titani K. , Terawaki Y. 
Published: October 2006

SummaryThe gene encoding cytotoxin (ctx) was cloned from Pseudomonas aeruginosa 158 and the nucleotide sequence was determined. The structural gene of ctx encodes the procytotoxin of 286 amino acid residues with a molecular mass of 31681 Daltons. Procytotoxin was activated by removal of 20 amino acid residues from the C terminus with trypsin. The cloned ctx gene was not expressed in either an Escherichia coli strain or a cytotoxin non‐producing strain of P. aeruginosa. An expression system for the ctx gene was constructed by placing the structural gene of ctx downstream of tac promoter on a broad host‐range vector plasmid.
Genetic factors influencing lignin peroxidase activity in Phanerochaete chrysosporium ME446
Authors: Raeder U. , Thompson W. , Broda P. 
Published: October 2006

SummaryHaploid recombinant progeny of Phanerochaete chrysosporium ME446, genome compositions of which had been defined by RFLP‐mapping, vary in their idiophasic behaviour. This allowed us to formulate a model of the sequence of idiophasic activities. One component of this variation, the amount of lignin peroxidase activity, is independent of the allele distributions of the lignin peroxidase gene clusters, but correlates with the allele distribution of another locus. This locus appears to control the spread of the lignin peroxidase‐active state within the idiophasic mycelial mat and may be the mating‐type locus. The successful determination of linkage relied on analysis of chromosome intervals rather than linkage to single markers; this approach should be generally useful for analysing quantitative characters by RFLP mapping.
A Mycoplasma genetic element resembling prokaryotic insertion sequences
Authors: Ferrell R. V. , Heidari M. B. , Wise K. S. , McIntosh M. A. 
Published: October 2006

SummaryNucleotide sequence analysis of two Mycoplasma hyopneumoniae‐derived copies of a repetitive genetic element revealed structural similarities to typical prokaryotic insertion sequences. This is the first such sequence identified in the class Mollicutes. The element spans approximately 1550bp, with 28bp inverted terminal repeats. Two open reading frames occur within the sequence, one potentially encoding a protein with a size‐variant alpha‐helical domain containing heptameric leucine periodicity. Hybridization data with several strains from each of two mycoplasma species showed that the repetitive sequence is variably distributed within the M. hyopneumoniae and Mycoplasma hyorhinis chromosomes and indicated that in some cases the repeated sequence is contained within a larger genetic element which may be the result of phage or plasmid Insertion.
Analysis of the chromosomal location of two copies of a Bordetella pertussis insertion sequence
Authors: McPheat W. L. , Hanson J. H. , Livey I. , Robertson J. S. 
Published: October 2006

SummaryIS481v1 and IS481v2 are two copies of a Bordetella pertussis insertion sequence element. We have shown that IS481V1 is located within 3kbp of the start of the adenylate cyclase gene whilst IS481v2 is immediately adjacent to the end of the agglutinogen 2 gene and provides the stop codon for that gene. In addition, IS481v1 and IS481v2 were present at these two specific sites in nine strains of B. pertussis, including two Phase IV strains which expressed neither adenylate cyclase nor agglutinogen 2 and three Phase I strains which did not express agglutinogen 2. The loss of expression in these strains is not the result of DNA rearrangements at the sites of IS481 v1 or IS481v2.
Molecular characterization of the proU loci of Salmonella typhimurium and Escherichia coli encoding osmoregulated glycine betaine transport systems
Authors: Stirling D. A. , Hulton C. S. J. , Waddell L. , Park S. F. , Stewart G. S. A. B. , Booth I. R. , Higgins C. F. 
Published: October 2006

SummaryThe proU loci of Salmonella typhimurium and Escherichia coli encode high‐affinity glycine betaine transport systems which play an important role in survival under osmotic stress. Transcription of the proU locus is tightly regulated by osmolarity and this regulation appears to be mediated by osmotically induced changes in DNA supercoiling. In order to study the regulatory mechanisms involved we have cloned and characterized the proU locus of S. typhimurium by an in vivo transductional procedure. The locus is shown to consist of at least three genes, designated proVWX, cotranscribed as a single operon. The first gene in the operon encodes a protein sharing considerable sequence identity with ATP‐binding proteins from other periplasmic transport systems. Unexpectedly, the highly expressed periplasmic glycine betaine binding protein was found to be encoded by a distal gene, proX, in the operon. The operon has no significant internal promoters but is expressed from a single osmoregulated promoter whose transcription start site has been mapped. The proU promoter of E. coli has also been sequenced and the transcription start site shown to be similar to that of S. typhimurium. Evidence is presented which suggests that, besides de novo glycine betaine uptake, an important function of ProU may be the recapture and recycling of other osmolytes that leak from the cell.
Transcriptional polarity enhances the contribution of the internal promoter, ilvEp, in the expression of the ilvGMEDA operon in wild‐type Escherichia coli K12
Authors: Lopes J. M. , Soliman N. , Smith P. K. , Lawther R. P. 
Published: October 2006

SummaryThe ilvG gene of Escherichia coli K12 produces a cryptic peptide as a result of a frameshift mutation located approximately halfway through the coding sequence of the gene. This mutation is polar on expression of the downstream genes (ilvEDA) because transcription terminates within the translationally barren region that results from the mutation. Contrary to this, Salmonella typhimurium produces a full‐length functional ilvG protein and is therefore unlikely to manifest this polarity event. E. coli K12 strains with mutations either in the ilvG gene (which restores a full‐length protein) or in the ρ gene, relieve this polarity suggesting that this event couples transcription and translation in a manner analogous to attenuation. This paper describes experiments designed to determine the molecular nature and location of the polarity event. Most significantly, this work establishes the contribution of the internal promoter (ilvEp, located downstream of the polar site) to the expression of the downstream genes in E. coli K12 wild‐type and mutant strains (ilvG) and by extension to the role of this promoter in S. typhimurium. This analysis suggests that ilvEp contributes as much as 90% of ilvEDA expression in wild‐type E. coli K12 and only 15% In wild‐type S. typhimurium when grown under non‐repressing conditions.
Localization and functional analysis of the regulated promoter from the Streptomyces glaucescens mel operon
Authors: Geistlich M. , Irniger S. , Hütter R. 
Published: October 2006

SummaryThe transcription initiation site of the meloperon from Streptomyces glaucescens, determined by S1 mapping and primer elongation experiments, lies 32 to 34bp upstream of the translation initiation codon of the first open reading frame. A total of 172 to 219bp upstream of the transcription start point are necessary for a fully active and regulated mel promoter. Deletion analysis, gel retardation assays and DNAse I footprint experiments facilitated division of the promoter into three functional domains, which include the RNA polymerase recognition site up to nucleotides ‐33 to ‐42, the binding region of a protein of assumed regulatory function between nucleotides ‐65 and ‐93, and an upstream activator site, located between positions ‐158 and ‐219.