摘要:

SummaryOpacity (Opa) proteins are a family of antigenically variable outer‐membrane proteins ofNeisseria meningitidis.Even among clonally related epidemic meningococcal isolates, there is greater variation of Opa protein expression than can be accounted for by theopagene repertoire of any individual strain. We characterized theopagenes of eight closely related Isolates of serogroup AN. meningitidis(subgroup IV‐1) from a recent meningitis epidemic in West Africa. DNA sequence analysis and Southern blot experiments indicated that changes occurred in theopagenes of these bacteria as they spread through the human population, over a relatively short period of time. Such changes in one or a few loci within a clonal population are referred to as microevolution. The distribution of sequences present in hypervariable (HV) regions of theopagenes suggests that duplication of all or part ofopagenes into otheropaloci changed the repertoire of Opa proteins that could be expressed. Additional variability in this gene family appears to have been introduced by horizontal exchange ofopasequences from other meningococcal strains and fromNeisseria gonorrhoeae.These results indicate that processes of recombination and genetic exchange contributed to variability in major surface antigens of this clonal population of pathogenic bacte

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01006.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryInduciblecatgenes from Gram‐positive bacteria are regulated by translation attenuation. The inducer chloramphenicol stalls a ribosome at a specific site in the leader ofcattranscripts; this destabillzes a downstream stem‐loop structure that normally sequesters the ribosome‐binding site for thecatstructural gene. The five‐amino‐acid peptide MVKTD that is synthesized when a ribosome has translated to the leader induction site is an inhibitor of peptidyl transferaseIn vitro.Thus, the peptide may be thein vivodeterminant of the site of ribosome stalling. Here we provide evidence that the leader pentapeptide can exert acis‐effect on its translating ribosomeIn vivo.Converting leader codon 6 to the ochre codon results in expression ofcat‐86in the absence of Inducer. We term this autoinduction. Autoinduction is abolished by mutations that change the amino‐acid sequence of the leader peptide but have no, or little, effect on the sequence of nucleotides at the leader stall site. In contrast, four nucleotide changes within the leader site occupied by the stalled ribosome that result in synonymous codon replacements do not diminish autoinduction. Our evidence indicates that thecat‐86leader pentapeptide can alter the function of its tra

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01007.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryYersinia enterocoliticais a facultative intracellular parasite, displaying the ability to grow saprophytically or invade and persist intracellularly in the mammalian reticuloendothelial system. The transition between such diverse environments requires the co‐ordinated regulation of specific sets of genes on both the chromosome and virulence plasmid. Temperature has a profound pleiotropic effect on gene expression and phenotypically promotes alterations in cell morphology, outer‐membrane protein synthesis, urease production, lipopolysaccharide synthesis, motility, and synthesis of genes involved in invasion of euKaryotic host cells. By examining thermoregulated flagella biosynthesis, we have determined that motility is repressed at 25° C (permissive temperature) with subinhibitory concentrations of novobiocin. These conditions also induce virulence gene expression suggesting novobiocin addition stimulates, at least partially, a high‐temperature environment. Furthermore, temperature‐shift experiments, usingY. enterocoliticacontaining pACYC184 as a reporter plasmid, indicate that thermo‐induced alterations of DNA supercoiling coincide with temperature‐induced phenotypic changes. A class of putative DNA gyrase mutant (novobiocin resistant) likewise demonstrates the 37° C phenotype when cultured at 25°C; it is non‐motile, urease negative, calcium growth dependent, and positive for Yop expression. These results support a model implicating DNA topology as a contributing factor ofY. enterocolitica

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01008.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryEra is an essential protein inEscherichia coliwhich binds both GTP and GDP and has an intrinsic GTPase activity. Studies on the role of GTP/GDP binding and GTPase activity in an attempt to understand its function lead to the observation that Era is autophosphorylated. The autophosphorylated reaction is specific for GTP and cannot use ATP as a phosphoryl group donor. The reaction velocity is of first order with respect to protein concentration, suggesting an intramolecular mechanism. Autophosphorylation occurs at serine and threonine residues. The major phosphorylated tryptic peptide isolated after autophosphorylation has been identified as ISITSR, from residue 33 to 38. The peptide contains the site of phosphorylation and two potential sites for serine and threonine phosphorylation. Subsequently, both the threonine residue at position 36 and the serine residue at position 37 were altered to alanine. The double mutant Era, but not individual single mutants, was unable to functionally complement the growth of anE. colistrain which cannot produce wild‐type Era protein at high temperature. This suggests that either threonine 36 or serine 37 has to exist for the function of EraIn vivo.phosphorylation of Era was also examined by two‐dimensional gel electrophoresis. Era has been previously assigned two distinct positions having two different X‐Y co‐ordinates: one of the spots (H032.0) was identified as phosphorylated Era, indicating that a substantial portion of Era in the cell is indeed phosphorylated. Therefore, Era autophosphorylation is likely to play an important physiological role in the cell. The sequence encoding theC‐terminus previously published had a missing C between A900 and GgO1. As a resuit of the frameshift, Era consists of 301 residues, 15 fewer than originaiiy

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01009.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryThe genome ofPhanerochaete chrysosporiumstrain ME446 contains multiple, non‐allelic, cellobiohydrolase I (CSHI)‐like sequences, at least two of which are expressed in a cellulose‐dependent manner. Each of the expressed genes contains two identically positioned introns within its coding region. The lengths and sequences of these introns are different and one is not excised from all transcripts, raising the possibility that subtly different protein products may be expressed from a common gene. Introns are also present upstream of both genes but these differ in number and position, as well as sequence and length. Endoglucanase‐like sequences could not be identified and it is suggested that variant CBHI‐like proteins may provide endoglucanase activity in th

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01010.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryIn order to understand how TEM‐1 β‐lactamase substrate specificity can be altered by mutation, amino acid residues 161 through to 170 were randomly mutagenized to sample all possible amino acid substitutions. The 161–170 region includes a portion of an omega loop structure, which is involved in the formation of the active‐site pocket. The percentage of random sequences that provide bacterial resistance to either ampicillin or to the extended‐spectrum cephalosporin ceftazidime was determined. It was found that the sequence requirements for wild‐type levels of ampicillin resistance are much more stringent than the sequence requirements for ceftazidime resistance. Surprisingly, more than 50% of all amino acid substitutions in the 161‐170 region result in levels of ceftazidime resistance at least three times greater than wild type. In addition, by increasing the level of the selection for ceftazidime resistance, substitutions that result in a greater than 100‐fold increase in ceftazidime resistance were identified. Characterization of altered β‐lactamase enzymes indicated that while their catalytic efficiency (Kcat/Km) for ceftazidime hydrolysis is higher, the enzymes are poorly expressed relative to wild‐t

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01011.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryReplacement of the CRP‐binding site of the gal control region by curved sequences can lead to the restoration of promoter strengthin vivo.One curved sequence called 5A6A, however, failed to do so. The genehnsexerts a strong negative control on the resulting 5A6Agalpromoter as well as on the distantblapromoter, specifically in a 5A6Agalcontext. The product of this gene, H‐NS, displays a better affinity for this particular insert compared to other curved sequences. Mechanisms by which H‐NS may repress promoters both at short and long distances from a favoured binding site are disc

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01012.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryXer site‐specific recombination is required for the stable inheritance of multicopy plasmids and the normal segregation of the bacterial chromosome inEscherichia coli.Two related recombinases and two accessory proteins are essential for Xer‐mediated recombination atcer, a recombination site in the plasmid ColE1 The accessory proteins, ArgR and PepA, function in ensuring that the Xer recombination reaction acts exclusively intramolecularly, converting plasmid dimers into monomers and not vice versa. PepA is an amino‐exopeptidase, but its molecular role in the Xer recombination mechanism is unclear. Here we show that a mutation directed at the presumptive active site of PepA creates a protein with no detectable peptidase activityin vitroorin vivo, but which still functions normality in Xer site‐specific recombinatio

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01013.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryMolecular analyses were applied to the genes encoding variants of the serosubtyping antigen, the class 1outer membrane protein (PorA), from 55 serogroup A Neisseria meningitidis strains. These genes were evolutionarily stable and exhibited a limited range of genetic variation, primarily generated by recombination. Translation of the gene sequences revealed a total of 19 distinct amino acid sequences in the variable regions of the protein, 6 of which were not recognized by currently available serosubtyping monoclonal antibodies. Knowledge of these aminoacid sequences permitted a rational re‐assignment of serosubtype names. Comparison of the complete genes with porA gene sequences from serogroup B and C meningococci showed that serogroup A possessed a limited number of the possibleporAgenes from a globally distributed gene pool. Each serogroup A subgroup was characterized by one of fourporAgene types, probably acquired upon subgroup divergence, which was stable over periods of decades and during epidemiological spread. Comparison with other variable genes (pil and iga) indicated that the three alleles were independently assorted within the subgroup, suggesting that their gene types were older than the subgroups in which they occurre

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01014.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryExpression of thevirBgene, the transcriptional regulator for the invasion genes encoded by the large plasmid ofShigella flexneri, is temperature‐regulated.virBtranscription is under the control of VirF and H‐NS, which act as positive and negative regulators, respectively, and is highly responsive to changes in DNA superhelicity. To further investigate the molecular mechanisms underlying the thermoregulation ofvirBtranscription, a mutant which expressed an invasion phenotype at both 30° C and 37° C was isolated using miniTn10‐Kan (miniKAN) random insertion mutagenesis. The insertion site was mapped to therhogene, and resulted in the addition of 11 amino acids to the C‐terminus of the Rho protein. Consequently, decreased transcription termination activity at a p‐dependent terminator, λtL1, was observed, in therhomutant, both the transcription ofvirBand expression of invasion genes were activated at 30°C and were less responsive to changes in temperature. The deregulation ofvirBexpression by the mutation was dependent upon thevirBpromoler, since the effects of the mutation onvirBtranscription were abolished when its promoler region was replaced by thetacpromoler. Temperature‐responsive changes in DNA topology, as determined by linking numbers of a reporter plasmid, showed that changes in DNA superhelicity in therhomutant were smaller than that in the wild type. Furthermore, when the mutant was grown in medium containing novobiocin, an inhibitor of DNA gyrase,virBtranscription at 30° C as well as at 37°C was greatly diminished. These results indicated that Rho protein could have a profound effect on topological temperature‐dependent changes in DNA structure, thus contributing to thermoregulation

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb01015.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY