bluuebunny/crossref_metadata_2025_split huggingface dataset:
Physical and genetic map of the Methanococcus voltae chromosome
Authors: Sitzmann J. , Klein A. 
Published: October 2006

SummaryA physical map of the Methanococcus voltae chromosome was constructed on the basis of restriction mapping and cross‐hybridization experiments, employing total and partial digests obtained with rarely cutting restriction enzymes. On the basis of the sum of the fragment sizes of digests with seven enzymes the chromosome length was calculated to be approximately 1900kb. The derived map is circular. Hybridization of gene probes to mapped restriction fragments has led to a genetic map of genes for structural RNAs as well as proteins, including enzymes involved in the methanogenic pathway.
Control of maltase synthesis in yeast
Authors: Needleman R. 
Published: October 2006

SummaryMaltose fermentation in Saccharomyces species requires the presence of at least one of five unlinked MAL loci: MAL1, MAL2, MAL3, MAL4 and MAL6. Each MAL locus is complex consisting of at least three genes: a trans‐acting activator, a maltose permease, and maltase. All the MAL loci show homology to each other both at the sequence level as determined by Southern transfer analysis and at the functional level as determined by complementation. We describe the organization of the MAL loci in yeast and the basic features of their regulation. The analysis of MAL has contributed to our understanding of the evolution of multigenic families. The global integration of carbohydrate metabolism, and gene regulation.
Molecular cloning, characterization and analysis of the regulation of the AR02 gene, encoding chorismate synthase, of Saccharomyces cerevisiae
Authors: Jones D. G. L. , Reusser U. , Braus G. H. 
Published: October 2006

SummaryWe describe here the cloning, characterization and analysis of the regulation of the ARO2 gene of Saccharomyces cerevisiae, the first reported study of a eukaryotic gene encoding chorismate synthase (E.C. 4.6.1.4). The gene contains an ORF of 1128 bp, encoding a protein with a calculated molecular mass of 40.8 kDa. ARO2 is regulated under the ‘general control system’ for amino acid biosynthesis by the transcriptional activator GCN4 which binds in vitro at three sites within the AR02 promoter.The AR02 gene product is highly similar to its Escherichia coli counterpart, with a 47% identity distributed over the entire length of the peptide. We therefore suggest that the S. cerevisiae chorismate synthase, in contrast to the Neurospora crassa enzyme, but like other chorismate synthases, is a monofunctional peptide, solely possessing chorismate synthase activity.
Characterization of kdgR, a gene of Erwinia chrysanthemi that regulates pectin degradation
Authors: Reverchon S. , Nasser W. , Robert‐Baudouy J. 
Published: October 2006

SummaryErwinia chrysanthemi is a phytopathogenic enterobacterium able to degrade the pectic fraction of plant cell walls. The kdgR negative regulatory gene controls all the genes involved in pectin catabolism, including the pel genes encoding pectate lyases. The E. chrysanthemi kdgR regulatory gene was subcloned in Escherichia coli where it was shown to be functional, since it repressed the expression of a pelE::uidA fusion. The nucleotide sequence of kdgR contained an open reading frame of 918bp preceded by classical transcriptional initiation signals. KdgR shows similarity to two other regulatory proteins, namely GylR, encoding an activator protein of the glycerol operon in Streptomyces coelicolor, and IclR, encoding a repressor of the acetate operon in Salmonella typhimurium and in Escherichia coli. Previously, comparison of regulatory regions of several genes controlled by kdgR revealed the existence of a conserved region which was proposed as a KdgR‐binding site. The 25 bp oligonucleotide AAAAAAGAAACATTG‐TTTCATTTGT corresponding to this consensus was substituted to the lac operator, at the beginning of transcription of the lacZ gene. This construct functioned as an operator for binding of the KdgR protein in vivo.
Pleiotropic control of Listeria monocytogenes virulence factors by a gene that is autoregulated
Authors: Mengaud J. , Dramsi S. , Gouin E. , Vazquez‐Boland J. A. , Milon G. , Cossart P. 
Published: October 2006

SummaryEvidence for ptelotropic activation of virulence genes in Listeria monocytogenes is presented. A complementation study of a spontaneous prfA‐deletion mutant and analysis of cassette and transposon insertion mutants showed that the gene prfA activates the transcription of four independent genes which code for a phosphatidyl‐inositol‐specific phospholipase C (gene plcA), listeriolysin O (gene hlyA), a metallo‐protease (gene prtA) and a lecithinase (gene prtC). Transcription of prfA is not constitutive. During the growth phase, two peaks of prfA transcript accumulation were observed: the first was during exponential growth, and the second was at the beginning of the stationary phase. In addition, two prf4‐specific transcripts of 2.2 kb and 1 kb are detected. Early in exponential growth, prfA is co‐transcribed with plcA which lies upstream prfA, giving rise to the 2.2 kb plcA‐prfA transcript. In late‐exponential growth and at the beginning of the stationary phase, prfA transcripts of 1 kb are predominantly detected. Our results demonstrate that since prfA controls plcA transcription, it also regulates its own synthesis.
Genetic mapping of starch‐ and Lambda‐receptor sites in maltoporin: identification of substitutions causing direct and indirect effects on binding sites by cysteine mutagenesis
Authors: Francis G. , Brennan L. , Stretton S. , Ferenci T. 
Published: October 2006

SummaryCysteine mutagenesis was used to test the proximity of 16 residues to protein‐ligand interaction sites in maltoporin (LamB protein). LamB protein with additional cysteines was incorporated into the outer membrane of Escherichia coli except with a Ser‐30→ Cys substitution. Phage Lambda and starch binding was assayed before and after incubation of mutants with six thiol‐specific reagents. Four categories of mutation were recognized on the basis of phenotype and modification for each of the Lambda‐ and starch binding sites. The thiol modification experiments helped to clarify whether the phenotype of a mutation was due to a substitution at the binding site or an indirect perturbation of the structure. This study suggests that the cysteine mutagenesis/thiol modification approach may be usefully applied to the operational mapping of surface‐accessible binding sites or epitopes.
Glucose repression in the yeast Saccharomyces cerevisiae
Authors: Trumbly R. J. 
Published: October 2006

SummaryUnderstanding the mechanism of glucose repression in yeast has proved to be a difficult and challenging problem. A multitude of genes in different pathways are repressed by glucose at the level of transcription. The SUC2 gene, which encodes invertase, is an excellent reporter gene for glucose repression, since its expression is controlled exclusively by this pathway. Genetic analysis has identified numerous regulatory mutations which can either prevent derepression of SUC2 or render its expression insensitive to glucose repression. These mutations allow us to sketch the outlines of a pathway for general glucose repression, which has several key elements: hexo‐kinase PII, encoded by HXK2, which seems to play a role in the sensing of glucose levels; the protein kinase encoded by SNF1, whose activity is required for derepression of many glucose‐repressible genes; and the MIG1 repressor protein, which binds to the upstream regions of SUC2 and other glucose‐repressible genes. Repression by MIG1 requires the activity of the CYC8 and TUP1 proteins. Glucose repression of other sets of genes seems to be controlled by the general glucose repression pathway acting in concert with other mechanisms. In the cases of the GAL genes and possibly CYC1, regulation is mediated by a cascade in which the general pathway represses expression of a positive transcriptional activator.
Isolation and analysis of IS6120, a new insertion sequence from Mycobacterium smegmatis
Authors: Guilhot C. , Gicquel B. , Davies J. , Martin C. 
Published: October 2006

SummaryInsertion sequence IS6120 from Mycobacterium smegmatis was identified by its ability to transpose into different sites in the lambda repressor gene, d857, carried on an Escherichia coli/mycobacteria shuttle plasmid. IS6120 is a novel 1.5 kb insertion sequence, which has 24‐bp imperfect terminal inverted repeats and generates 9‐bp duplications of the target DNA following insertion. IS6120 is present in at least three copies in M. smegmatis but was not found in other species, including Mycobacterium tuberculosis. Nucleotide sequence analysis revealed that IS6120 contains two open reading frames, one of which encodes a putative transposase with similarities to those found in IS256 from Staphylococcus aureus, IST2 from Thiobacillus ferrooxidans, and ISRm3 from Rhizobium meliloti. The fact that IS6120 does not recognize a consensus target sequence for insertion and has no homologous sequences in the other strains studied makes IS6120 useful for transposon mutagenesis in mycobacteria.
Outer membranes of Gram‐negative bacteria are permeable to steroid probes
Authors: Plésiat Patrick , Nikaido Hiroshi 
Published: October 2006

SummaryThe permeability of bacterial outer membranes was assayed by coupling the influx of highly hydrophobic probes, 3‐oxosteroids, with their subsequent oxidation catalysed by 3‐oxosteroid δ1‐dehydrogenase, expressed from a gene cloned from Pseudomonas testosteroni. In Salmonella typhimurium producing wild‐type lipopolysaccharide, the permeability coefficients for uncharged steroids were 0.45 to 1 × 10‐5 cm S‐1 and the diffusion appeared to occur mainly through the lipid bilayer domains of the outer membrane. These rates are one or two magnitudes lower than that expected for their diffusion through the usual biological membranes. The permeation rates were markedly increased (up to 100 times) when the lipopolysaccharide leaflet was perturbed either by adding deacylpolymyxin or by introducing mutations leading to the production of deep rough lipopolysaccharides. An amphiphilic, negatively charged probe, testosterone hemisuccinate, penetrated much more slowly than the uncharged steroids. Study of various Gram‐negative species revealed that P. testosteroni, Pseudomonas acidovorans, and Acinetobacter calcoaceticus showed higher outer membrane permeability to steroid probes and higher susceptibility to hydrophobic agents such as fusidic acid, novobiocin and crystal violet relative to S. typhimurium and Escherichia coli.
Characterization of the Escherichia coli codBA operon encoding cytosine permease and cytosine deaminase
Authors: Danielsen S. , Kilstrup M. , Barilla K. , Jochimsen B. , Neuhard J. 
Published: October 2006

SummaryThe nucleotide sequence of a 3.1 kb segment carrying the cytosjne deaminase gene (codA) from Esctierichia coli was determined. The sequence revealed the presence of two open reading frames, the first (codB) specifying a highly hydrophobic polypeptide and the second specifying cytosine deaminase. A two‐codon overlap between the two reading frames indicates that they constitute an operon. Transcription of the operon was found to be regulated by exogenous purines. Polypeptides specified by each of the two reading frames were expressed in minicells, and the codB gene product was found to be highly enriched in the membrane fraction. Uptake experiments showed that the CodB protein is required for cytosine transport into the cell and that the intracellular accumulation of cytosine correlated with the codB gene dose. A topological model for the cytosine permease in the cytoplasmic membrane is proposed.
Nucleotide sequence analysis and serologic characterization of the Mycobacterium intracellulare homologue of the Mycobacterium tuberculosis 19 kDa antigen
Authors: Nair Jaygopal , Rouse David A. , Morris Sheldon L. 
Published: October 2006

SummaryDisseminated Mycobacterium avium/Mycobacterium intracellulare complex (MAC) disease is a frequent complication in patients with the acquired immune deficiency syndrome (AIDS). In this report, we present the nucleotide sequence of the M. intracellulare MI22 gene. Computer sequence comparisons reveal that the MI22 gene, which encodes a serologically active protein, has 78% DNA sequence identity and 77% protein sequence identity with the seroreactive 19 kDa Mycobacterium tuberculosis lipoprotein antigen. Southern blot hybridizations indicate that an MI22 gene probe binds similar—‐sized restriction fragments in M. tuberculosis and M. intracellulare genomic DNA. In addition, immunoblot analyses demonstrate that MI22 is recognized by sera from tuberculosis patients. These data further support the existence of 19 kDa MAC and M. tuberculosis protein homologues. Phase partitioning experiments and the presence of a consensus lipid modification site in the deduced MI22 protein sequence strongly suggest that MI22 is also a lipoprotein. Comparative analyses of these mycobacterial antigenic homologues may provide the basis for the design of species‐specific diagnostic reagents.
Identification of individual amino acids required for secretion within the haemolysin (HlyA) C‐terminal targeting region
Authors: Kenny Brendan , Taylor Sally , Holland I. Barry 
Published: October 2006

SummaryThe release of haemolysin from Escherichia coli involves direct secretion across both the inner and outer membranes. Secretion of HlyA is dependent upon a specific membrane export complex composed of HlyB, ‐D and possibly TolC. HlyA is targeted to the medium via the membrane translocation complex, by a novel C‐terminal secretion signal. Previous studies involving deletion and fusion analyses have given contradictory results for the minimal length (20–60 residues) of this HlyA signal region and little is known of the nature of the specific residues and structural features required for function.In this study we have analysed, quantitatively, the effect upon secretion of many point mutations introduced into the HlyA C‐terminus. The results indicate the presence of a minimal secretion signal domain whose proximal boundary extends to at least residue –46 and which contains at least four Individual residues essential for maximal secretion levels. We propose that such residues act co‐operatively, forming multiple contact points with the translocator proteins, with the ‘best fit’ promoting maximal levels of secretion.
Identification and characterization of a novel insertion sequence, IS1106, downstream of the porA gene in B15 Neisseria meningitidis
Authors: Knight A. I. , Ni H. , Cartwright K. A. V. , McFadden J. J. 
Published: October 2006

SummaryExamination of Neisseria meningitidis strains associated with endemic meningococcal disease demonstrated differences in the number of copies of a repetitive sequence. Characterization of a copy of this repetitive sequence present in B15 strains has revealed the presence of a novel insertion sequence (S11106) located within a complex repetitive region downstream of the gene for the major surface antigen (porA). IS1106 has a length of 1137 bp and is flanked by 36bp inverted repeats. Two open reading frames (0RF1 and 0RF2) are present in opposite strands in codon–codon register with 0RF2 entirely located within ORF1. The predicted protein from 0RF1 demonstrates homology with the 5A protein of IS5 (Kroger and Hobom, 1962). Strains from two independent outbreaks of B15 meningococcal disease in the UK were found to contain the same genomic deletion removing a copy of IS1106 downstream of the porA gene.
Amino acid sequences of several Bacillus subtilis proteins modified by apparent guanylylation
Authors: Mitchell Clayton , Morris Paul W. , Vary James C. 
Published: October 2006

Summary Bacillus subtilis cell extracts, prepared at different times during growth, contained several proteins that were apparently guanylylated in vitro with [α‐32P]‐GTP. Four of the proteins were partially purified and the N‐terminal amino acid sequences (13 to 20 residues) were determined. One sequence had 84% identity to Bacillus stearothermophlius triosephosphate isomerase, two were 100% identical to the predicted sequences of the B. subtilis ptsI and ptsH genes while no identity was found for the fourth sequence. This apparent guanylylation occurred with proteins involved in glucose metabolism, although the significance is unknown.
The role of the pro‐sequence in the processing and secretion of the thermolysin‐like neutral protease from Bacillus cereus
Authors: Wetmore Diana R. , Wong Sui‐Lam , Roche Rodney S. 
Published: October 2006

SummaryThe Bacillus cereus cnp gene coding for the thermolysin‐like neutral protease (TNP) has been cloned, sequenced, and expressed in Bacillus subtilis. The protease is first produced as a pre‐pro‐protein (Mr= 61000); the pro‐peptide is approximately two‐thirds of the size of the mature protein. The pro‐sequence has been compared with those of six other TNPs, and significant homologies have been found. Additionally, the TNP pro‐sequences are shown to be homologous to the pro‐sequence of Pseudomonas aeruginosa elastase. A mutant has been constructed from cnp, in which 23 amino acids upstream from the pro‐protein processing site have been deleted. This region has no homologous analogue in any of the other TNP pro‐sequences. The detection results in a delay of six to eight hours in detection of active protease in the growth medium, as well as a 75% decrease in maximum protease production. N‐terminal analysis of the mutant mature protein demonstrates that the processing site is unaltered by the pro‐sequence deletion. The deletion must, therefore, modulate the kinetics of processing and/or secretion of the pro‐protein.
Characterization of transformation‐deficient mutants of Acinetobacter calcoaceticus
Authors: Palmen Ronald , Vosman Ben , Kok Ruben , van der Zee J. Rob , Hellingwerf Klaas J. 
Published: October 2006

SummaryThree Acinetobacter calcoaceticus transformation‐deficient mutants, obtained by insertional mutagenesis with the nptII gene, have been characterized physiologically. One mutant (AAC211) was found to be completely transformation deficient, while two others, AAC213 and AAC214, were severely impaired in transformation efficiency (100–1000 times lower than the wild type). The latter applied to both chromosomal as well as plasmid DNA. Analysis of the chromosomal DNA fragments flanking the nptll gene in the mutants showed that mutants AAC213 and AAC214 had an insertion of the nptll gene in the same chromosomal region, but that they were the result of two independent mutational events, whereas the insertion in mutant AAC211 was at a different position. None of the three mutants showed phenotypic or genotypic characteristics typical of a RecA‐deficient strain.
NtcA, a global nitrogen regulator from the cyanobacterium Synechococcus that belongs to the Crp family of bacterial regulators
Authors: Vega‐Palas M. A. , Flores E. , Herrero A. 
Published: October 2006

SummaryThe gene ntcA is required for full expression of proteins subject to ammonium repression in the cyanobacterium Synechococcus. A 3.1 kb DNA fragment able to complement an ntcA mutant was digested with exonuclease III, and deleted fragments of different sizes were tested for complementation of that mutant, allowing the localization of its mutation within a BamHI‐HindIII genomic fragment of c. 0.4 kb. Insertion of a chloramphenicol‐resistance‐encoding gene cassette into both the BamHI and the HindiIII sites of wild‐type Synechococcus resulted in a pleiotropic, nitrogen‐assimilation‐minus phenotype, corroborating the presence of the ntcA gene in that genomic region. Sequencing of DNA in this region showed the presence of an open reading frame that included both the BamHI and the HindIII sites. The ntcA gene product, NtcA, is a protein of 24817 Da which belongs to a family of bacterial transcriptional activators that, among others, includes Crp and Fnr from Escherichia coli. Of special biological significance, it appears, is the presence of a conserved helix‐turn‐helix motif in the sequence close to the C‐terminal end of all the proteins in the family. The gene ntcA is proposed to encode a transcriptional activator of genes subject to nitrogen control in Synechococcus.
Identification and characterization of narQ, a second nitrate sensor for nitrate‐dependent gene regulation in Escherichia coli
Authors: Chiang Robin C. , Cavicchioli Ricardo , Gunsalus Robert P. 
Published: October 2006

SummaryIn response to nitrate availability, Escherichia coli regulates the synthesis of a number of enzymes involved in anaerobic respiration and fermentation. When nitrate is present, nitrate reductase (narGHJI) gene expression is induced, while expression of the DMSO/TMAO reductase (dmsABC, fumarate reductase (frdABCD)and fermentation related genes are repressed. The narL and narX gene products are required for this nitrate‐dependent control, and apparently function as members of a two‐component regulatory system. NarX is a presumed sensor‐transmitter for nitrate and possibly molybdenum detection. The presumed response‐regulator, NarL, when activated by NarX then binds at the regulatory DNA sites of genes to modulate their expression. In this study a third nitrate regulatory gene, narQ, was identified that also participates in nitrate‐dependent gene regulation. Strains defective in either narQ or narX alone exhibited no nitrate‐dependent phenotype whereas mutants defective in both narQ and narX were fully inactive for nitrate‐dependent repression or activation. In all conditions tested, this regulation required a functional narL gene product. These findings suggest that the narX and narQ products have complementary sensor‐transmitter functions for nitrate detection, and can work independently to activate NarL, for eliciting nitrate‐dependent regulation of anaerobic electron transport and fermentation functions. The narQ gene was cloned, sequenced, and compared with the narX gene. Both gene products are similar in size, hydrophobicity, and sequence, and contain a highly conserved histidine residue common to sensor–transmitter proteins.
NADH formation by Na+‐coupled reversed electron transfer in Klebsiella pneumoniae
Authors: Dan Pfenninger‐Li Xiao , Dimroth Peter 
Published: October 2006

SummaryCitrate is fermented by Klebsiella pneumoniae to 2 acetate, 0.5 formate and 1.2 CO2. The formation of <1 formate and >1 CO2 per citrate can be accounted for by the oxidation of formate to CO2 in order to provide reducing equivalents for the assimilation of citrate into cell carbon. A membrane‐bound electron transport chain is apparently involved in NADH synthesis by these cells. The electrons from formate oxidation to CO2 are used to reduce ubiquinone to ubiquinol by membrane‐bound formate dehydrogenase and ubiquinol further delivers its electrons to NAD+, if this endergonic reaction is powered by δμTNa+. The endogenous NADH level of K. pneumoniae cells thus increased in the presence of formate in response to a δpNa+ >‐100 mV. NADH formation was completely abolished in the presence of oxygen or after addition of hydroxyquinoline‐N‐oxide, a specific inhibitor of the Na+‐translocating NADH:ubiqurnone oxidoreductase. The increase of endogenous NADH was dependent on the δpNa+ applied to the cells. Inverted membrane vesicles of K. pneumoniae catalysed the reduction of NAD+ to NADH with formate as electron donor after application of delta;μTNa+ of about 120 mV consisting of δpNa+ of 60 mV and ΔΨ of the same magnitude. Neither the δpNa+ nor the ΔΨ of this size alone was sufficient to drive the endergonic reaction. Strictly anaerobic conditions were required for NADH formation and hydroxyquinoline‐N‐oxide completely inactivated the reaction. It is suggested that NADH formation by reversed δμTNa+‐coupled electron transfer in these cells is an essential requirement for the synthesis of cell material from citrate.