bluuebunny/crossref_metadata_2025_split huggingface dataset:
Two linked genes for outer membrane proteins are absent in four non‐disease strains ofHaemophilus somnus
Authors: Cole S. P. , Guiney D. G. , Corbeil L. B. 
Published: October 2006

SummaryLinked genes encoding two outer membrane proteins (p76 and a family of proteins, p120) of the bovine pathogen,Haemophilus somnus, were investigated. The p120 group was previously shown to have immunoglobulin‐binding activity and to react with polyclonal antiserum specific for a 270 kDa antigen (p270) which also had immunoglobulin Fc‐binding activity. By Western blotting we showed that the p76 antigen also reacted with this antiserum. The p270, p120, and p76 antigens were undetectable in four serum‐sensitive isolates from asymptomatic carriers but were present in the two serum‐resistant virulent strains tested. Genes for p120 and p76 were subcloned on non‐overlapping pUC plasmids from a cosmid (pHS1) originally cloned from a serum‐resistant strain. InEscheriehia coli, plasmid pHS138 expressed p76, while the p120 antigens were produced by pHS140. Southern blots of DNA from the above six strains ofH. somnususing probes derived from pHS1 subclones showed that a 13.4kb sequence was missing from the four serum‐sensitive strains, but not the two serum‐resistant strains. This segment included most of the insert in pHS138 and all of the pHS140 insert. The data indicate that p76 and the p120 proteins are absent from serum‐sensitive strains because the coding sequences are missing, raising the possibility of insertion of these genes into the chromosome of both serum‐resistant strains, or deletion from the four serum‐sensitive strains.
The divergent promoters mediating transcription of the par locus of plasmid RP4 are subject to autoregulation
Authors: Eberl Leo , Givskov Michael , Schwab Helmut 
Published: October 2006

SummaryThe partitioning region of broad‐host‐range plasmid RP4 contains four genes (parA, parB, parC, and parD) that encode products essential for partition activity. Two divergently arranged promoters located in the rntercistronrc region between parC and parD mediate transcription of these genes. The transcriptional initiation sites for both promoters were determined by primer extension. Transcriptional fusions were used to show that parA, parB, and parCare combined in an operon, while parD constitutes a separate transcription unit. Both parCBA (genes in order of transcription) and parD are negatively autoregulated at the level of transcription by the gene products of parA and parD, respectively. parD promoter mutants which have become insensitive to repression by parD were isolated. Comparison of wild type and the mutant parD promoter sequences indicated that three short repeats are likely involved in the negative regulation of this promoter. Potentially these sequence elements comprise target sites for the ParD protein.
Isolation of arginine auxotrophs, cloning by mutant complementation, and sequence analysis of the argC gene from the cyanobacterium Anabaena species PCC 7120
Authors: Floriano B. , Herrero A. , Flores E. 
Published: October 2006

SummaryArginine auxotrophs of the dinitrogen‐fixing cyanobacterium Anabaena species strain PCC 7120 were isolated after ultraviolet light mutagenesis and penicillin enrichment. Two of these auxotrophs were complemented by a cosmid gene library of the wild‐type strain established in Escherichia coli that was transferred en masse to the mutants by conjugation. The gene complementing one of those mutants was found to complement an E. coli argC mutant. Sequencing analysis of the gene showed that it encodes a 322‐residue polypeptide that is homologous to the ArgC protein of E. coli, Bacillus subtilis and Streptomyces clavuligerus and to the C‐terminal moiety of the Sac‐charomyces cerevisiae ARG5,6 gene product, N‐acetylglutamate semialdehyde dehydrogenase. A cysteine residue present in a highly conserved domain in the five proteins is probably located in the active site of the enzyme. Conserved among the ArgC proteins, sequences resembling the primary structure of nucleotide‐binding domains are also found. Downstream of the Anabaena argC gene seven nearly perfect repeats of a heptanucleotide (consensus sequence: 5′‐CTAATGA‐3′) are found.
The ω‐endotoxin protein family displays a hydrophobic motif that might be implicated in toxicity
Authors: Pereyra‐Alférez B. , Bravo A. , Quintero R. , Soberón X. 
Published: October 2006

SummaryA computer‐based analysis of hydropathy and surface probability of representative members of each class of the Cry family of proteins was performed. A highly conserved hydrophobic motif within the previously described block, D2, is present not only In lepidopteran toxin genes but also in toxins active against diptera and coleoptera. An interesting feature of this hydrophobic motif is the presence of an aspartic residue (highly hydrophilic) in its middle part. Comparison with the amino acid sequence from diphtheria toxin showed that it also contains a hydrophobic motif similar to the one present in the Bacillus thuringiensis toxins. It also contains an aspartic residue in the middle part and some speculations are presented on the function of this specific region with regard to the toxic mechanism of action.
Molecular cloning and characterization of two lincomycin‐resistance genes, ImrA and ImrB, from Streptomyces lincolnensls 78–11
Authors: Zhang Hui‐Zhan , Schmidt Heike , Piepersberg Wolfgang 
Published: October 2006

SummaryTwo different lincomycin‐resistance determinants (ImrA and ImrB) from Streptomyces lincolnensis 78–11 were cloned in Streptomyces lividans 66 TK23. The gene ImrA was localized on a 2.16 kb fragment, the determined nucleotide sequence of which encoded a single open reading frame 1446 bp long. Analysis of the deduced amino acid sequence suggested the presence of 12 membrane‐spanning domains and showed significant similarities to the methylenomycin‐resistance protein (Mmr) from Streptomyces coelicolor, the QacA protein from Staphylococcus aureus, and several tetracycline‐resistance proteins from both Gram‐positive and Gram‐negative bacteria, as well as to some sugar‐transport proteins from Escherichia coli. The ImrB gene was actively expressed from a 2.7 kb fragment. An open reading frame of 837 bp could be localized which encoded a protein that was significantly similar to 23S rRNA adenine(2058)‐N‐methyltransferases conferring macrolide—lincosamide—streptogramin resistance. LmrB also had putative rRNA methyltransferase activity since lincomycin resistance of ribosomes was induced in ImrB‐containing strains. Surprisingly, both enzymes, LmrA and LmrB, had a substrate specificity restricted to lincomycin and did not cause resistance to other lincosamides such as celesticetin and clindamycin, or to macrolides.
Isolation and characterization of an ndvB locus from Rhizobium fredii
Authors: Bhagwat Arvind A. , Tully Raymond E. , Keister Donald L. 
Published: October 2006

SummaryA gene (ndvB) in Rhizobium meliloti that is essential for nodule development in Medicago sativa (alfalfa), specifies synthesis of a large membrane protein. This protein appears to be an intermediate in β‐1, 2‐glucan synthesis by the microsymbiont. Southern hybridization analysis showed strong homology between an ndvB (chvB) probe and genomic DNA of R. fredii but not from Bradyrhizobium japonicum. A cosmid clone containing the putative ndvB locus was isolated from a Rhizobium fredii gene library. The cosmid clone which complemented R. meliloti ndvB mutants for synthesis of β‐1, 2‐glucans and effective nodulation of alfalfa was mapped and subcloned. Fragment‐specific Tn5 mutagenesis followed by homologous recombination into the R. fredii genome indicated that the region was essential for β‐1, 2‐glucan synthesis and for formation of an effective symbiosis with Glycine max (soybean).
Mutagenesis induced by bacterial UmuDC proteins and their plasmid homologues
Authors: Woodgate Roger , Sedgwick Steven G. 
Published: October 2006

SummaryThe popular image of a world full of pollutants mutating DNA is only partly true since there are relatively few agents which can subtly and directly change base coding; for example, some alkylating agents alter guanine so that it pairs like adenine. Many more mutagens are less subtle and simply destroy coding altogether rather than changing it. Such mutagens include ultraviolet light, X‐rays, DNA cross‐linkers and other agents which make DNA breaks or large adducts. In Escherichia coli, mutagenesis by these agents occurs during a DNA repair process which increases cell survival but with an inherent possibility of changing the original sequence. Such mutagenic DNA repair is, in part, encoded by the E. coli umuDC operon. This article reviews the structure, function, regulation and evolution of the umuDC operon and similar genes found both in other species and on naturally occurring plasmids.
Ethanol‐induced and glucose‐insensitive alcohol dehydrogenase activity in the yeast Kluyveromyces lactis
Authors: Mazzoni Cristina , Saliola Michele , Falcone Claudio 
Published: October 2006

SummaryThe alcohol dehydrogenase (ADH) system in the yeast Kluyveromyces lactis is encoded by four ADH genes. In this paper we report evidence that at least three of these genes are transcribed and transcribed into protein. KIADH1 and KIADH2, which encode cytoplasmic activities, are preferentially expressed in glucose‐grown cells with respect to ethanol‐grown cells. KIADH4, which encodes one of the two activities localized within mitochondria, is induced at the transcriptional level in the presence of ethanol as is the ADH2 gene in Saccharomyces cerevlslae. However the regulation of the expression of the K. lactis gene is completely different from that of ADH2 and of other known ADH genes in that KIADH4 is insensitive to glucose repression and is not expressed on non‐fermentabie carbon sources other than ethanol. This km6 of regulation can be clearly observed in non‐fermenting strains, where the induction of KIADH4 is dependent on the addition of ethanol to the medium. On the contrary, in fermenting strains KIADH4 is always induced by ethanol or acetaldehyde produced endocellularly and this results in constitutive expression of the gene aiso in the presence of glucose. The mitochondrial localization of the activity encoded by KIADH4 and the peculiar regulation of this gene could be related to the fact that K. lactis is a petite negative yeast in which some mitochondrial functions seem to be essential for cell viability.
Analysis of the regulation of the pelBC genes in Erwinia chrysanthemi 3937
Authors: Hugouvieux‐Cotte‐Pattat Nicole , Robert‐Baudouy Janine 
Published: October 2006

Summary Erwinia chrysanthemi secretes five major isoenzymes of pectate lyases encoded by the pelABCDE genes. The nucleotide sequence of the region surrounding the pelB gene of E. chrysanthemi 3937 was determined, including the regulatory regions involved in pelB and pelC expression. Analysis of the transcripts showed that transcription of pelB or pelC gave, in both cases, only one transcript. The transcription initiation sites of both pelB and pelC were precisely determined as well as the position of the transcription termination of pelB. The pelB and pelC promoters are very similar, showing a good homology with the ‐35 consensus region but tow homology with the ‐10 consensus. In both cases a KdgR‐box overlaps the ‐35 region. The pelC gene may have two KdgR operators. Moreover, the pelB and pelC genes are preceded by other sequences presenting the typical symmetry of operator sites that could be involved in more specific regulations. Comparison of E. chrysanthemi pel regulatory regions revealed three classes of homology: pelA, pelB—pelC and pelD—pelE. The sole regulatory sequence conserved among the three classes corresponds to the KdgR‐binding site. Moreover, all the pel regulatory regions are AT‐rich in contrast to the coding regions which are GC‐rich. Gel retardation experiments with fragments overlapping the pelB or pelC regulatory regions demonstrated that the KdgR protein specifically binds to these regions. Other proteins probably also interact with these DNA fragments. Transcription of pelB terminates in a region corresponding to a GC‐rich inverted repeat followed by a run of T residues, typical of rho‐independent transcription termination sites. Moreover, preliminary results imply that a region adjacent to pelC provoke, directly or indirectly, the repression of pelB and pelC expression.
gltF, a member of the gltBDF operon of Escherichia coli, is involved in nitrogen‐regulated gene expression
Authors: Castaño Irene , Flores Noemi , Valle Fernando , Covarrubias Alejandra A. , Bolivar Francisco 
Published: October 2006

SummaryWe report here the construction and analysis of insertional mutations in each of the three genes of the gltBDF operon and the nucleotide sequence of the region downstream from gltD. Two open reading frames were identified, the first of which corresponds to gltF. The gltB and gltD genes code for the large and small subunits, respectively, of the enzyme glutamate synthase (GOGAT). gltF codes for a protein, with a molecular mass of 26350Da, which is required for Ntr induction. Histidase synthesis was determined as a measure of Ntr function. First, insertions in gltB, gltD or gltF all prevent Ntr induction. Second, complementation analysis indicates that high‐level expression of both the gltD and gltF genes is required for the induction of the Ntr enzymes under nitrogen‐limiting conditions, indicating that the phenotype of the gltB insertion probably results from polarity on gltD and gltF. Third, glutamate‐dependent repression of the glt operon appears to be mediated by the product of the gltF gene. Thus, the gltBDF operon of Escherichia coli is involved in induction of the so‐called Ntr enzymes in response to nitrogen deprivation, as well as in glutamate biosynthesis.
Surface exclusion specificity of the TraT lipoprotein is determined by single alterations in a five‐amino‐acid region of the protein
Authors: Harrison Jacqueline L. , Taylor Ian M. , Platt Karen , O'Connor C. David 
Published: October 2006

SummaryThe TraT protein is a highly cell‐surface‐exposed lipoprotein specified by F‐like plasmids that confers serum resistance and blocks the conjugative transfer of plasmids to cells bearing identical or closely related plasmids, a process known as surface exclusion. The TraT protein specified by the antibiotic‐resistance plasmid R6‐5 was purified to apparent homogeneity. When added to mating mixtures, TraT blocked the transfer of plasmids belonging to Surface Exclusion Group IV (Sfx IV) but had no significant effect on the transfer of plasmids belonging to other groups. Additionally, the purified protein had a protective effect on bacterial cells incubated in serum, indicating that it does not have to be located on the cell surface to mediate serum resistance. To localize regions of the protein that were responsible for surface exclusion specificity, the amino acid sequence of the TraT protein specified by ColB2‐K98 (Sfx II) was determined by cloning and sequencing of the corresponding gene. Comparison of the derived sequence with those of the F and R100‐1 proteins indicated that surface exclusion specificity of TraT is determined by single alterations in a five‐amino‐acid region (residues 116–120). This was confirmed by segment swapping experiments in which the specificity of the R6‐5 TraT protein (Sfx IV) was switched to that of the COIB2‐K98 protein (Sfx II). Our results suggest that the region defined by residues 116–120 is located on the external face of the outer membrane and interacts specifically with the donor cell in surface exclusion.
Gene cassettes from the insert region of integrons are excised as covalently closed circles
Authors: Collis Christina M. , Hall Ruth M. 
Published: October 2006

SummaryIntegrons are DNA elements which generally include one or more discrete gene cassettes inserted at a specific site. We have recently proposed a model for the acquisition and dissemination of genes found in the insert region of integrons, which requires the existence of circularized gene cassettes. Evidence for the existence of covalently closed circular molecules consisting of one or more gene cassettes has now been obtained. Low levels of small molecules which hybridize to probes specific for individual gene cassettes were detected in plasmid DNA isolated from cells containing a plasmid which includes an integron fragment with three gene cassettes aacC1, orfE and aadA2. These molecules were only detected when the gene encoding the integron DNA integrase was also present and are thus products of site‐specific cassette excision. The excised cassettes have been shown to be in the form of covalently closed supercoiled circles, by digestion with restriction enzymes exonuclease III and DNase I. The circular excision products detected included either one cassette, aadA2 or orfE, two cassettes, aacC1 and orfE or all three cassettes. The predicted sequence of the recombinant junction in the excised aadA2 cassette confirmed that excision was precise. The predicted unique sequences of the 59‐base elements associated with individual genes in the circular cassette form were compiled, and the sequences of the seven‐base core sites which flank 59‐base elements are now, with few exceptions, exact inverted repeats.
Mycobacterial protein antigens: a compilation
Authors: Young D. B. , Kaufmann S. H. E. , Hermans P. W. M. , Thole J. E. R. 
Published: October 2006

SummaryIn response to recommendations from the Steering Committees responsible for co‐ordination of World Health Organization programmes for research on the immunology of leprosy (IMMLEP) and tuberculosis (IMMTUB), a list was prepared summarizing the properties of mycobacterial proteins currently under investigation with respect to their immunological activities. After consultation with more than 40 laboratories world‐wide this list was extended to form the compilation shown below and is intended to provide a comprehensive and convenient reference for future studies in this field.
Molecular and immunological analysis of a fibronectin‐binding protein antigen secreted by Mycobacterium leprae
Authors: Thole J. E. R. , Schöningh R. , Janson A. A. M. , Garbe T. , Cornelisse Y. E. , Clark‐Curtiss J. E. , Kolk A. H. J. , Ottenhoff T. H. M. , De Vries R. R. P. , Abou‐Zeid C. 
Published: October 2006

SummaryBy screening a Mycobacterium lepraeλgt11 genomic DNA library with leprosy‐patient sera we have previously identified 50 recombinant clones that expressed novel M. leprae antigens (Sathish et al., 1990). In this study, we show by DNA sequencing and immunoblot analysis that three of these clones express a M. leprae homologue of the fibronectin‐binding antigen 85 complex of mycobacteria. The complete gene was characterized and it encodes a 327‐amino‐acid polypeptide, consisting of a consensus signal sequence of 38 amino acids followed by a mature protein of 289 amino acids. This is the first sequence of a member of the M. leprae antigen 85 complex, and Southern blotting analysis indicated the presence of multiple genes of the 85 complex in the genome of M. leprae. The amino acid sequence displays 75–85% sequence identity with components of the antigen 85 complex from M. tuberculosis, M. bovis BCG and M. kansasii. Furthermore, antibodies to the antigen 85 complex of M. tuberculosis and M. bovis BCG reacted with two fusion proteins containing the amino acid regions 55–266 and 265–327 of the M. leprae protein. The M. leprae 30/31 kDa protein induces strong humoral and cellular responses, as judged by Western blot analysis with patient sera and proliferation of T cells derived from healthy individuals and leprosy patients. Amino acid regions 55–266 and 265–327 both were shown to bind to fibronectin, indicating the presence of at least two fibronectin‐binding sites on the M. leprae protein. These data indicate that this 30/31 kDa protein is not only important in the immune response against M. leprae, but may also have a biological role in the interaction of this bacillus with the human host.
Isolation, characterization, and genetic analysis of monosomic, aneuploid mutants of Candida albicans
Authors: Barton R. C. , Gull K. 
Published: October 2006

SummaryA white, prototrophic Candida albicans strain, heterozygous for the ADE2 gene (ade2/ADE2), was treated with the antimitotic agent methyl benzimidazole carbamate, and yielded red, adenine‐requiring colonies at a rate of 4 × 10‐3, an order of magnitude higher than the spontaneous rate of Ade‐ colony formation. These red Ade‐ colonies were small, growing at approximately half the rate of the parent strain, and gave rise to large red colonies spontaneously. When the chromosomes of the small red colonies were separated by pulsed‐field gel electrophoresis, the band hybridizing with the ADE2 gene was diminished in staining intensity by half relative to the parent and large red‐colony strains. Restriction fragment‐length polymorphism analysis and auxotrophic mutant spectra after mutagenesis suggested that the small red Ade‐ strains were monosomic aneuploids lacking one of a pair of chromosome homologues, while the large red strains had regained a homologue, presumably via a second non‐disjunction event. Parasexual genetic analysis of two of the auxotrophs isolated from a putative aneuploid suggested that both mutations were linked to the ADE2 gene. These experiments suggest that targeted chromosome loss and monosomic, aneuploid strains have the potential to extend the scope of genetic analysis in this diploid, asexual organism.
Cloning and characterization of a phospholipase gene from Erwinia chrysanthemi EC16.
Authors: Keen N. T. , Ridgway D. , Boyd C. 
Published: October 2006

SummaryA single gene (plcA) was cloned from a cosmid library of Erwinia chrysanthemi EC16 DNA that encoded an extracellular phospholipase. The gene was subcloned and DNA sequence data showed the presence of a single open reading frame encoding a protein with a predicted size of 39kDa. The coding region was G+C‐rich and the protein had a predicted basic isoelectric point. The protein showed no significant homology with others in the PIR library, including other phospholipases. When overexpressed in Escherichia coli cells, the plcA gene directed production of a c. 39kDa protein that was largely localized in the periplasm, but its N‐terminal amino acid sequence was that of the native protein predicted from DNA sequence data. Unlike the wild‐type bacterium, an E. chrysanthemi EC16 marker exchange mutant of the plcA gene did not secrete extracellular phospholipase activity into the medium. However, no detectable change was observed in terms of the virulence of the mutant strain on potato tubers or chrysanthemum stems.
The relationship between disulphide bond formation, processing and secretion of lipo‐β‐lactamase in yeast
Authors: Shani O. , Pines O. 
Published: October 2006

SummaryThe hybrid prokaryotic lipo‐β‐lactamase mature and precursor proteins spontaneously form an intramolecular disulphide bond when oxidized in vitro. When expressed in Saccharomyces cerevisiae (in vivo) the lipo‐β‐lactamase precursor is in a reduced form whereas the majority of the mature protein is oxidized. The results indicate that in yeast, the lipo‐β‐lactamase precursor is first processed (the signal peptide is removed) and then oxidized to form a disulphide bond in the mature protein. Reduced‐mature lipo‐β‐lactamase was found to reach the yeast periplasm and the process depends on endoplasmic reticulum (ER) entry even though the protein is not oxidized. This result is remarkable since in eukaryotes, disulphide bond formation occurs in the ER. Oxidized mature lipo‐β‐lactamase can also be released from the sphaeroplast into the yeast periplasm. Mutant lipo‐β‐lactamase genes in which cysteine residue 131 was changed to either tyrosine or threonine, were efficiently processed and secreted in yeast, which is consistent with the finding that reduced‐mature non‐mutant lipo‐β‐lactamase can be secreted. We discuss the possibility that the folding mechanism of lipo‐β‐lactamase in vitro may be fundamentally different from the process in the eukaryotic system of S. cerevisiae.