bluuebunny/crossref_metadata_2025_split huggingface dataset:
Translational repression of TEM β‐lactamase synthesisas a response of Escherichia coli to heat shock
Authors: Kuriki Y. 
Published: October 2006

SummaryAs a result of a temperature shift‐up from 30°C to 42°C, β‐lactamase synthesis in Escherichia coli carrying pBR322 ceased transiently, even though the level of β‐lactamase mRNA was not altered. pBR328‐directed pre‐β‐lactamase synthesis in an in vitro transcription‐translation coupled system was also repressed by incubation at the higher temperature. Translation of the lacZ sequence from the amp translation start signal, inserted into the open reading frame vector pORF1, was also repressed transiently upon the temperature shift‐up. Pre‐heating of the in vitro coupled system at 45°C specifically reduced its capacity for pre‐β‐lactamase synthesis. This capacity was restored by the addition of a 160000 × g supernatant prepared from E. coli grown at 30°C, but not by the supernatant from the cells incubated at 42°C. These and other results Indicate that (i) the 160000 × g supernatant contains a heat‐labile protein(s) that is required for efficient initiation of the translation of pre‐β‐lactamase mRNA, and (ii) the heat shock‐induced repression of β‐lactamase synthesis is due to inactivation of the protein(s) in the 160000 × g supernatant.
Characterization of amber mutations in bacteriophage Mu transposase: a functional analysis of the protein
Authors: Desmet L. , Faelen M. , Gama M.‐J. , Ferhat A. , Toussaint A. 
Published: October 2006

SummaryWe have characterized a series of amber mutations in the A gene of bacteriophage Mu encoding the phage transposase. We tested different activities of these mutant proteins either in a sup0 strain or in different sup bacteria. In conjunction with the results described in the accompanying paper by Bétermier et al. (1989) we find that the C‐terminus of the protein is not absolutely essential for global transposase function, but is essential for phage growth. Specific binding to Mu ends is defined by a more central domain. Our results also reinforce the previous findings (Bétermier et al., 1987) that more than one protein may be specified by the A gene.
Cloning a genomic region required for a high‐affinity iron‐uptake system in Rhizobium meliloti 1021
Authors: Gill P. R. , Neilands J. B. 
Published: October 2006

SummaryA collection of transposon‐induced mutants of Rhizobium meliloti 1021 defective in siderophore‐mediated iron assimilation were obtained and classified as biosynthetic, transport or regulatory. Several of the mutations were cloned and the adjacent sequences were used to acquire complementing DNA from the wild type. A single genomic region of about 35kb complemented all of the mutants deficient in production of the siderophore.
The transition state transcription regulator AbrB of Bacillus subtilis is autoregulated during vegetative growth
Authors: Strauch M. A. , Perego M. , Burbulys D. , Hoch J. A. 
Published: October 2006

SummaryThe DNA‐binding AbrB protein of Bacillus subtilis is an ambiactive transcriptional regulator of genes expressed during the transition state between vegetative growth and the onset of stationary phase and sporulation. Studies on the transcriptional control of AbrB synthesis using abrB‐lacZ fusions indicated that the abrB gene was autoregulated. This was consistent with the observation that purified AbrB protein bound specifically to the promoter region of its own gene in DNase I protection experiments. The structural gene mutation abrB4 abolished the autoregulatlon and purified AbrB4 protein did not have the promoter binding properties associated with the wild‐type protein. Both AbrB and AbrB4 proteins were shown to be hexamers of 10500 Dalton subunits and subunit exchange occurred between the proteins in vitro. However, the presence of only one or two mutant subunits dramaticaly altered the DNA‐binding ability of the multimeric protein. The results support a model in which autoregulation of the abrB gene is an important factor in preventing sporulation‐associated genes from being expressed during vegetative growth.
Expression of the recE gene during induction of the SOS response in Bacillus subtilis recombination‐deficient strains
Authors: Gassel M. , Alonso J. C. 
Published: October 2006

SummaryA transcriptional fusion of the recE gene to a reporter gene has been constructed. Expression of recE was found to be induced upon damage to DNA with either mitomycin C or nalidixic acid. This specific transcriptional induction is blocked by a recE mutation. Mutations affecting the recB, recF and recL gene products markedly reduced induction. However, de‐repression of recE seems to be independent of the ATP‐dependent DNase activity of the exonuclease V enzyme (also called AddAB enzyme).
Membrane topology of penicillin‐binding protein 3 of Escherichia coli
Authors: Bowler L. D. , Spratt B. G. 
Published: October 2006

SummaryThe β‐lactamase fusion vector, pJBS633, has been used to analyse the organization of penicillin‐binding protein 3 (PBP3) in the cytoplasmic membrane of Escherichia coli. The fusion junctions in 84 in‐frame fusions of the coding region of mature TEM β‐lactamase to random positions within the PBP3 gene were determined. Fusions of β‐lactamase to 61 different positions in PBP3 were obtained. Fusions to positions within the first 31 residues of PBP3 resulted in enzymatically active fusion proteins which could not protect single cells of E. coli from killing by ampicillin, indicating that the β‐lactamase moieties of these fusion proteins were not translocated to the peri‐plasm. However, all fusions that contained: ≥36 residues of PBP3 provided single ceils of E coli with substantial levels of resistance to ampicillin, indicating that the β‐lactamase moieties of these fusion proteins were translocated to the periplasm. PBP3 therefore appeared to have a simple membrane topology with residues 36 to the carboxy‐terminus exposed on the periplasmic side of the cytoplasmic membrane. This topology was confirmed by showing that PBP3 was protected from proteolytic digestion at the cytoplasmic side of the inner membrane but was completely digested by proteolytic attack from the periplasmic side. PBP3 was only inserted in the cytoplasmic membrane at its amino terminus since replacement of its putative lipoprotein signal peptide with a normal signal peptide resulted in a water‐soluble, periplasmic form of the enzyme. The periplasmic form of PBP3 retained its penicillin‐binding activity and appeared to be truly water‐soluble since it fractionated, in the absence of detergents, with the expected molecular weight on Sephadex G‐100 and was not retarded by hydrophobic interaction chromatography on Phenyl‐Superose.
The Bradyrhizobium japonicum fixBCX operon: identification of fixX and of a 5’mRNA region affecting the level of the fixSCX transcript
Authors: Gubler M. , Zürcher T. , Hennecke H. 
Published: October 2006

SummaryThe Bradyrhizobfum japonicum fixX gene was identified and shown to be essential for symbiotic and free‐living, microaerobic nitrogen fixation. The fixX gene encodes a ferredoxin‐like protein which may be Involved in a redox process (electron transport?) essential for nitrogenase activity. This gene was localized downstream of fixC and its expression was dependent on the fixB promoter, providing evidence for the existence of a fixBCX operon. Mutagenesis and sequence analysis of the unusually long, 709 bp leader region between the fixB promoter and the fixB structural gene did not reveal the presence of a nif or fix gene that was absolutely essential for nitrogen fixation. However, a short open reading frame (ORF) within this region encoding a polypeptide of 35 amino acids (ORF35) was shown to be efficiently translated. Chromosomal deletion of a 400bp DNA fragment covering ORF35 resulted in a three‐fold reduction of the fixSCX mRNA level, which in turn also reduced the nitrogen fixation activity of this mutant. This suggests a possible post‐transcriptional control mechanism for the expression of the fixBCX operon involving the stabilization of fixBCX mRNA by ribosomes actively translating ORF35.
Cloning of a surface protein antigen gene from serotype c Streptococcus mutans
Authors: Okahashi N. , Sasakawa C. , Yoshikawa M. , Hamada S. , Koga T. 
Published: October 2006

SummaryThe structural gene for a 190kD protein antigen (PAc) of Streptococcus mutans MT8148 (serotype c) was cloned into the plasmid vector pUC118. SDS‐polyacrylamide gel electrophoresis and Western immunoblotting showed that the Escherichia coli harbouring the chimaeric plasmid produced multiple polypeptides of 190‐210kD. Immunodiffusion analysis revealed that the cloned PAc had the same specific determinants as S. mutans PAc. The cloned pac gene was mapped, and its transcriptional orientation was determined by characterizing deletion mutants of the chimaeric plasmid. Southern blot analysis with the cloned gene sequence as a probe revealed the presence of a homologous sequence in DNAs from sero‐types e and f S. mutans. PAc‐defective mutants were constructed by inserting an erythromycin‐resistance gene into the pac gene. The cell‐surface hydrophobicity of the mutants was lower than that of the parent strain.
Mutations produced by DNA polymerase III holoenzyme of Escherichia coli after in vitro synthesis in the absence of single‐strand binding protein
Authors: Carraway M. , Rewinski C. , Marinus M. G. 
Published: October 2006

SummarySingle‐stranded plasmid DNA, containing the mnt gene, was replicated in vitro with DNA polymerase III holoenzyme. Escherichia colimutH bacteria, defective in mismatch repair, were transformed with the products of in vitro synthesis. Mutations in mnt were readily identified and 33 out of 65 isolates were single base changes including transition, transversion and frameshift mutations. The remaining 32 isolates were deletions of apparently random length and substitutions (deletion/insertions). The intergenic deletions as well as the transition and frameshift mutations were identical to those previously isolated from mismatch repair‐defective cells in vivo.
Geographically distinct isolates of Mycobacterium leprae exhibit no genotypic diversity by restriction fragment‐length polymorphism analysis
Authors: Williams D. L. , Gillis T. P. , Portaels F. 
Published: October 2006

SummaryDifferentiation of microorganisms for taxonomic purposes is based primarily on phenotypic characteristics, which are the direct or cumulative result of gene expression. Since expression of phenotypic characteristics usually relies on in vitro growth of a microorganism, non‐cultivable organisms, such as Mycobacterium leprae, present major problems for the identification of potential variants based on phenotypic similarities or differences between individual isolates. We have employed the use of restriction fragment‐length polymorphism (RFLP) analysis of chromosomal DNA of M. leprae isolates, including human isolates from geographically distinct regions of the world and isolates from a Sooty Mangabey monkey and an armadillo, to assess the relatedness among these isolates. Restriction endonuclease (Eco RI, Bst EII, Pst I, and Pvu I) digests of chromosomal DNA were analysed using DNA probes encoding all or part of the 12kD, 18kD, 28kD, 65kD and 70kD proteins of M. leprae as well as a probe containing an M. leprae‐specific sequence repeated up to 20 times in the M. leprae chromosome. Comparison of the resulting autoradiographs showed that the RFLP patterns were all identical, indicating that these isolates contained no polymorphism with respect to the restriction endonuclease sites analysed. In addition, RFLP patterns of two separate human M. leprae isolates remained unchanged after three cycles of experimental infection in the armadillo model. These results indicated that the M. leprae isolates tested in this study were indistinguishable at the genotypic level, strongly suggesting homogeneity among members of this species.
Functional importance of the Escherichia coli ribosomal RNA leader box A sequence for post‐transcriptional events
Authors: Theißen G. , Behrens S. E. , Wagner R. 
Published: October 2006

SummaryTo shed more light on the controversial findings concerning the functional participation of the highly conserved nut‐like leader box A sequence element in ribosomal RNA transcription antitermination we have carried out a mutational study. We have substituted the box A and combined this mutation with several deletions comprising the rRNA leader elements box B, box C and the tL region. The mutations are located within the genuine rrnB operon cloned on multicopy plasmids. We determined the effects of the mutations on cell growth, rRNA accumulation and ribosomal subunit stoichiometry. Cells transformed with the mutated plasmids were affected in their growth rate, and showed a surprising deficiency of the promoter‐proximal 16S compared to the 23S RNA, indicative of a post‐transcriptional degradation event. Accordingly, we could demonstrate a reduced amount of free 305 relative to 50S ribosomal subunits in exponentially growing cells. Similar stoichiometric aberrations in the ribosome pool were detected in conditionally Nus factor‐defective strains. The results show that the leader box A sequence within rRNA operons has important post‐transcriptional functions for 16S RNA stability and ribosomal subunit stoichiometry. A model is proposed, describing the biogenesis and quality control of ribosomes based on rRNA leader and Nus‐factor interactions. It is compatible with the previously observed effects of box A in antitermination.
Spore colour in Streptomyces coelicolor A3(2) involves the developmentally regulated synthesis of a compound biosynthetically related to polyketide antibiotics
Authors: Davis N. K. , Chater K. F. 
Published: October 2006

Summary Streptomyces coelicolor produces spores whose development of a grey colour requires the activity of the whiE locus. The cloned whiE locus was identified after mobilization into a whiE mutant of a library of S. coelicolor DNA inserted into a transmissible plasmid vector. The whiE region of the cloned DNA was localized both by subcloning and by mutagenesis of the cloned DNA with the Streptomyces transposon Tn4560. Nucleotide sequencing of this region revealed seven open reading frames, of which six show homo‐logy at the level of deduced gene products with genes involved in the synthesis of polyketide antibiotics. A previously described S. coelicolor DNA segment encoding biosynthesis of a brown pigment (Horinouchi and Beppu, 1985) corresponds to the cloned whiE DNA. It is proposed that whiE is normally expressed only in the aerial hyphae, where the biosynthetic product is responsible for spore colour.
Transcriptional analysis of the glnB‐glnA region of Rhizobium leguminosarum biovar viciae
Authors: Chiurazzi M. , Laccarino M. 
Published: October 2006

SummaryWe report that the glnB and gln A genes of Rhizobium leguminosarum biovar viciae are preceded by promoters located upstream of each gene. We find the presence of a glnB–glnA and a glnA mRNA whose intracellular concentration changes two‐ to three‐fold when R leguminosarum is grown on different nitrogen sources. Primer extension analysis shows unique transcriptional initiation sites upstream of glnB and glnA. The glnB promoter is rpoN(ntrA)‐dependent, while the glnA promoter does not contain a typical consensus sequence for previously described promoters.In Klebsiella pneumoniae the glnB promoter requires active ntrC and ntrA genes and a DNA fragment containing 53 nucleotides upstream of the transcription initiation site shows full promoter activity, thus indicating that no NtrC binding sites are necessary for this activation in the glnB upstream region.
Direction of DNA entry in competent cells of Bacillus subtilis
Authors: Vagner V. , Claverys J.‐P. , Ehrlich S. D. , Méjean V. 
Published: October 2006

SummaryDirection of DNA entry in Bacillus subtilis competent cells was studied using molecules in which only one of the two strands was radioactively labelled. The label was either distributed homogeneously or was localized in a small region of the strand, in the centre or at one of the ends. Regardless of the distribution and the position of the label, similar amounts of radioactivity were taken up by the cells exposed to the labelled molecules. This suggests that DNA enters B. subtilis either by two different uptake systems having opposite polarities, or by a single non‐polar system.
The yeast phosphatase system
Authors: Vogel K. , Hinnen A. 
Published: October 2006

SummaryYeast cells produce a set of enzymes which are involved in the metabolism of phosphate, and include acid and alkaline phosphatases as well as permeases. Most of these enzymes are synthesized in response to the presence or absence of inorganic phosphate. In the past few years a considerable amount of genetic and molecular evidence has accumulated and a rather precise overall picture emerges which describes the mechanism of phosphate control at the level of gene activation. This mini‐review summarizes these data. The main focus lies on the regulatory features associated with the control of transcription of PH05, a gene coding for most of the regulated acid phosphatase activity produced by yeast cells.
TonB and the Gram‐negative dilemma
Authors: Postle K. 
Published: October 2006

SummaryTonB protein serves as an energy transducer to couple cytoplasmic membrane energy to high‐affinity active transport of iron siderophores and vitamin B12 across the outer membranes of Gram‐negative bacteria. The biochemical mechanism of the energy transduction remains to be determined, but important details are already known. TonB is targeted to and anchored in the cytoplasmic membrane by a single membrane‐spanning domain and spans the periplasm to physically interact with outer‐membrane receptors of the transport ligands. TonB‐dependent energy transduction is modulated by ExbB protein, which stabilizes TonB, and possibly by several other proteins including ExbC, ExbD, and TolQ. TonB has a relatively short functional half‐life that is accelerated when rates of active transport across the outer membrane are increased. A model that incorporates this information, as well as some tempered speculation, is presented.
Vitamin B12 transport in Escherichia coli: energy coupling between membranes
Authors: Kadner R. J. 
Published: October 2006

SummaryCells of Escherichia coli possess high‐affinity active transport systems of vitamin B12 and iron‐siderophore complexes. Specific outer‐membrane proteins carry out the energy‐dependent transport across the outer membrane, in conjunction with the TonB coupling protein. Mutagenesis experiments have identified a conserved region near the amino‐terminus of the outer‐membrane transporters that is necessary for energy‐coupled transport. The ability of extragenic suppressor mutations in tonB to correct the transport defect indicates that TonB couples the proton‐motive force to the outer‐membrane proteins by direct contact.
Efflux‐mediated antiseptic resistance gene qacA from Staphylococcus aureus: common ancestry with tetracycline‐ and sugar‐transport proteins
Authors: Rouch D. A. , Cram D. S. , Di Berardino D. , Littlejohn T. G. , Skurray R. A. 
Published: October 2006

SummaryResistance to intercalating dyes (ethidium, acriflavine) and other organic cations, such as quaternary ammonium‐type antiseptic compounds, mediated by the Staphylococcus aureus ptasmid pSK1 is specified by an energy‐dependent export mechanism encoded by the qacA gene. From nucleotide sequence analysis, qacA is predicted to encode a protein of Mr 55017 containing 514 amino acids. The gene is likely to initiate with a CUG codon, and a 36bp palindrome immediately preceding qacA, along with an upstream reading frame with homology to the TetR repressors, may be components of a regulatory circuit. The putative polypeptide specified by qacA has properties typical of a cytoplasmic membrane protein, and is indicated to be a member of a transport protein family that includes proteins reponsible for export‐mediated resistance to tetracycline and methylenomycin, and uptake of sugars and quinate. The analysis suggests that N‐ and C‐terminal regions of these proteins are involved in energy coupling (proton translocation) and substrate transport, respectively. The last common ancestor of the qacA and related tet (tetracycline resistance) lineages is inferred to have been repressor controlled, as occurs for modern tet determinants from Gram‐negative, but not those from Gram‐positive, bacteria.
Isolated DNA repeat region from fcrA76, the Fc‐binding protein gene from an M‐type 76 strain of group A streptococci, encodes a protein with Fc‐binding activity
Authors: Heath D. G. , Boyle M. D. P. , Cleary P. P. 
Published: October 2006

SummaryThe DNA repeat region of fcrA76, the gene encoding a group A streptococcal Fc‐binding protein, was sub‐cloned in‐frame into an Escherichia coli plasmid expression vector. The expressed protein product displayed the same Fc‐binding properties as the full‐length Fc‐binding protein expressed from fcrA76. The affinity‐purified, full‐length Fc‐binding protein was found to compete with staphylococcal protein A or streptococcal protein G for binding to beads coated with human IgG. These results are consistent with earlier studies suggesting that the binding sites on human IgG for protein A, protein G and the type II Fc‐binding protein from group A streptococci are located at the interface of the CH2 and CH3 domains of the Fc region.
Activation of a cryptic crystal protein gene of Bacillus thuringiensis subspecies kurstaki by gene fusion and determination of the crystal protein insecticidal specificity
Authors: Dankocsik C. , Donovan W. P. , Jany C. S. 
Published: October 2006

SummaryDNA hybridization with the insecticidal crystal protein gene cryIIA (formerly cryBI) of Bacillus thuringiensis supspecies kurstaki has shown that subspecies kurstaki contains a cryIIA‐related sequence in addition to the cryIIA gene (Donovan et al., 1988a). We have cloned the cryIIA‐related sequence and have determined that the sequence, which has been designated cryIIB, is 89% identical to the cryIIA gene. Recombinant B. thuringiensis cells harbouring the cloned cryIIB gene produced very little CryIIB protein. A high level of production of the CryIIB protein was achieved by fusing the regulatory region of the cryIIIA crystal protein gene to the cryIIB gene. The CryIIB protein was found to be highly toxic to Lymantria dispar, Heliothis virescens and Trichoplusia ni, and was not toxic to Aedes aegypti.
RNase E, an endoribonuclease, has a general role in the chemical decay of Escherichia coli mRNA: evidence that rne and ams are the same genetic locus
Authors: Mudd E. A. , Krisch H. M. , Higgins C. F. 
Published: October 2006

Summary Escherichia coli RNase E is known to process RNA precursors at specific sites. We show that this endoribonuclease has a general role in E. coli mRNA turnover and affects the stability of specific transcripts. The effect of the rne mutation on functional stability of mRNA was much less pronounced than that on chemical stability, although the expression of some genes was affected. The E. coli ams (altered mRNA stability) mutation was found to have phenotypes indistinguishable from those of the rne mutation, affecting both 9S RNA and T4 gene 32 mRNA processing. The rne and ams mutations were both complemented by the same 3.7kb fragment of E. coli DNA and are probably allelic. RNase E is the first endoribonuclease Identified as having a general role in the chemical decay of E. coli mRNA.