摘要:

In one experimental system, several handles on the molecular mechanism of apparent adaptive mutation have emerged. The system is reversion of alacframe‐shift mutation inEscherichia coli. The molecular handles include a requirement for homologous recombination; the implication of DNA double‐strand breaks as a molecular intermediate; a unique sequence spectrum of −1 deletions in mononucleotide repeats which implies polymerase errors, and also implies a failure of post‐synthesis mismatch repair on those errors; and the involvement of sexual functions at some stage of the process. These molecular handles are revealing an unexpected new mechanism of muta

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020185.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

Coliphage λ employs systems of transcription termination and antitermination to regulate gene expression. Early gene expression is regulated by the phage‐encoded N protein working with a series ofEscherichia coliproteins, Nus, at RNA sites, NUT, to modify RNA polymerase to a termination‐resistant form. Expression of λ late genes is regulated by the phage‐encoded Q antitermination protein. Q, which appears to use only one host factor, acts at a DNA site,qut, to modify RNA polymerase to a termination‐resistant form. This review focuses on recent studies which show that: (i) N can mediate antiterminationin vitro, independent of Nus proteins, (ii) Early genes in another lambdoid phage HK022 are also regulated by antitermination, where only an RNA signal appears necessary and sufficient to create a termination‐resistant RNA polymerase. (iii) A part of thequtsignal appears to be read from the non‐template DNA strand. (iv) A host‐encoded inhibitor of N antitermination appears to act through the NUT site as well as with the α subunit of RNA polymerase, and is antagonized

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020191.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

A virulence‐associated region in the genome ofDichelobacter nodosushas been shown to contain an integrase gene which is highly related to the integrases ofShigella flexneriphage Sf6 and coliphages P4 and φR73, together with open reading frames (vapB, CandD) related to genes borne on plasmids inNeisseria gonorrhoeae, Escherichia coli, Actinobacillus actinomycetemcomitansandTreponema denticola. Similar to P4 and φR73, thevapregion is bracketed by putative bacteriophageattsites and is adjacent to a tRNA gene, which suggests that thevapregion has been derived by the integration of a bacteriophage, or a plasmid carrying a bacteriophage‐related integrase gene. Many similarities in genes and genes clusters encoding virulence determinants have been found in distantly related bacteria. These genes are often located on plasmids in one organism but on the chromosome in others, implying that transmission of the genes has been followed by integration. Thus, the events which have generated thevapregions ofD. nodosusmay represent a common mechanism for transfer of virulence determinants. A number of genes involved in the virulence of bacterial pathogens are found on integrated bacteriophages, and we suggest that others will prove to be associated with tRNA genes and/or integrase genes derived from bacteriophages. The use of tRNA genes as integration sites for many bacteriophages and plasmids may favour intergeneric transmission, as tRNA genes are highly cons

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020201.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

Cloning of therfbgenes ofShigella flexneri2a intoEscherichia coliK‐12 strain DH1 results in the synthesis of lipopolysaccharides (LPS) with an O‐antigen chain having type antigen IV and group antigens 3,4. During genetic studies of theserfbgenes inE. coliK‐12, we observed that strains harbouring plasmids with certain mutations (inversion and transposon insertions) which should have blocked O‐antigen synthesis nevertheless still produced LPS with O‐antigen chains. These LPS migrated differently on silver‐stained SDS—polyacrylamide gels, compared with the LPS produced by wild‐typerfbgenes, and the group 3,4 antigens were barely detectable, suggesting that the O‐antigen was altered. Investigation of the genetic determinants for production of the altered O‐antigen/LPS indicated that: (i) these LPS are produced as a result of mutations which are either polar onrfbFor inactivaterfbF; (ii) therfbXgene product (or a similar protein in theE. coliK‐12rfbregion) is needed for production of the altered O‐antigen in the form of LPS; (iii) therfbGgene product is required for the production of both the parental and altered LPS; (iv) the dTDP‐rhamnose biosynthesis genes are required. Additionally, anE. coliK‐12 gene product(s) encoded outside therfbregion also contributes to production of the O‐antigen of the altered LPS. An antiserum raised to the altered LPS from strain DH1(pPM2217 (rfbX::Tn1725)) was found to cross‐react with nearly allS. flexneriserotypes, and with the altered LPS produced by other DH1 strains harbouring plasmids with differentrfbmutations, as described above. The reactivity of the altered LPS with a panel of monoclonal antibodies specific for variousS. flexneriO‐antigen type and group antigens demonstrated that their O‐antigen components were closely related to that ofS. flexneriserotype 4. The RfbF and RfbG proteins were shown to have similarity to rhamnose transferases, and we identified a motif common to the N‐termini of 6‐deoxy‐hexose nucleotide sugar transferases. We propose that theE. coliK‐12 strains harbouring the mutatedS. flexneri rfbgenes produce LPS with a hybrid O‐antigen as a consequence of inactivation of RfbF and complementation by anE. coliK‐12 gene product. Analysis of the genetic and immunochemical data suggested a possible structure

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020209.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

We report the discovery of novel subcellular structures related to bacterial nitrogen fixation in the strictly respiratory diazotrophic bacteriumAzoarcussp. BH72, which was isolated as an endophyte from Kallar grass. Nitrogenase is derepressed under microaerobic conditions at O2concentrations in the micromolar range. With increasing O2deprivation, bacteria can develop into a hyperinduced state, which is characterized by high specific rates of respiration and efficient nitrogen fixation at approximately 30 nM O2. Ultrastructural analysis of cells in the course of hyperinduction revealed that complex intracytoplasmic membrane systems are formed, which consist of stacks of membranes and which are absent under standard nitrogen‐fixing conditions. The iron protein of nitrogenase was highly enriched on these membranes, as evidenced by immunohistochemical studies. Membrane deficiency in NifH/K−mutants, a deletion mutant in thenifKgene and the character of NH4+‐grown cells suggested, in concert with the membrane localization of nitrogenase, that these structures are specialized membranes related to nitrogen fixation. We propose the term ‘diazosomes’ for them. Development of intracytoplasmic membranes coincides with the appearance of a high‐molecular‐mass form of the iron protein of nitrogenase, which was detectable in membrane fractions. Mutational analysis, and determination of the N‐terminal amino acid sequence indicate that thenifHgene product is covalently modified by a mechanism probably different from adenosine diphosphoribosylation. Development of diazosomes in nitrogen‐fixing cells can be induced in pure cultures and in co‐culture with a fungus isolated from the rhizosph

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020225.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

STBsecretion‐deficient mutants were isolated using the synthetic transposon TnβIaM. Cultures were plated using a double‐membrane system of cellulose acetate and nitrocellulose placed on Luria agar plates containing carbenicillin. The STBbound to the underlying nitrocellulose membrane was detected with anti‐STBantibodies. The altered genes of two STBsecretion‐deficient mutants were identified by conjugation and complementation astoICanddsbA. In cultures of well‐characterizeddsbAandtoICmutants, STBwas absent from the culture supernatant. The role ofToICand DsbA in the secretion of peptides is

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020237.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

HfIB, also called FtsH, is an essentialEscherichia coliprotein involved in the proteolysis of the heat‐shock regulator σ32and of the phage regulator λcll. ThehfIB1(Ts) allele (formerly calledftsH1) conferring temperature‐sensitive growth at 42°C is suppressed by loss of the ferric‐uptake repressor Fur and by anaerobic growth. We show here that suppression requires TonB‐dependent Fe(III) transport in thehfIB1(Ts)furmutant during aerobic growth at 42°C and Feo‐dependent Fe(II) transport during anaerobic growth at 42°C. Temperature‐resistant growth ofhfIB1(Ts) strains is also observed at 42°C in the presence of a high concentration of Fe(II), Ni(II), Mn(II) or Co(II) salts, but not in the presence of Zn(II), Cd(II), Cu(II), Mg(II), Ca(II) or Cr(III) salts. However, neither Ni(II) nor afurmutation permits growth in the complete absence of HfIB. The heat‐shock response, evaluated by anhtpG::lacZfusion, is overinduced inhfIB1(Ts) strains at 42°C because of stabilization of σ32. Growth in the presence of Ni(II) or in the absence of the Fur repressor abolishes this overinduction in thehfIB1(Ts) strain, and, in thehfIB1(Ts)furmutant, σ32is no longer stabilized at 42°C. These results reinforce the recent observation that HfIB is a metalloprotease active against σ32in vitroand suggest that it can associate functionallyin vivowith Fe(II), Ni(II)

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020247.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

TheospCgene was amplified by the polymerase chain reaction from each of 76 Lyme diseaseBorreliastrains. Restriction fragment length polymorphism (RFLP) analysis demonstrated 33 distinct RFLP types; two additional RFLP types were identified from publishedospCsequences. For each RFLP type, at least oneospCgene was sequenced and the degree of sequence relatedness examined by construction of anospCgene tree. The genes were extremely diverse, with sequence identity ranging from 74.4% to 99.0%; the majority of changes are localized within the central portion of the molecule. A comparison ofospCsequences suggests that recombination occurs frequently betweenospCalleles; this genetic exchange is proposed to be mediated by lateral transfer ofospCsequences. Evidence indicates that recombination occurs betweenospCgenes from the sameBorreliaspecies (i.e.B. afzeliiandB. garinii) as well as between differentBorreliaspecies (i.e.B. afzeliiandB. garinii, B. burgdorferiand genogroup DN127).

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020257.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

The broad‐host‐range plasmid pAMβ1 from Gram‐positive bacteria encodes a resolvase, designated Resβ, which shares homology with the proteins of the resolvase—invertase family. Here we report the purification andin vitrocharacterization of Resβ. This resolvase is particular in two aspects: it has an atypical binding site and requires a cofactor to promote resolutionin vitro. Resβ binds to two regions within its resolution siteres. One contains two inverted repeats (R1 and R2), the other contains only one repeat (R3). The cofactor required for resolutionin vitrois present in crude extracts of bothBacillus subtilisandEscherichia coliand can be substituted by theE. colihistone‐like protein HU. The possible mode of action of HU in the resolution process

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020271.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY

摘要:

Wall‐less prokaryotes in the genusMycoplasmainclude over 90 species of infectious agents whose pathogenicity for humans and other animals is currently being assessed. Molecular characterization of surface proteins is critical in this regard but is hampered by the lack of genetic systems in these organisms. We used TnphoAtransposition to systematically mutagenize, inEscherichia coli, a genomic plasmid library constructed fromMycoplasma fermentans, a potential human pathogen. The strategy circumvented problems of expressing mycoplasma genes containing UGA (Trp) codons and relied on the construction of the vector pG7ZCW, designed to reduce TnphoAtransposition into vector sequences. FunctionalphoAgene fusions directly identified genes encoding 19 putative membrane‐associated proteins ofM. fermentans. Sequences of fusion constructs defined three types of export sequence: (1) non‐cleavable, membrane‐spanning sequences, (2) signal peptides with signal peptidase (SPase) I‐like cleavage sites, and (3) signal peptides with SPase II‐like lipoprotein‐cleavage sites which, like most other mycoplasmal lipoprotein signals analysed to date, differed from those in several Gram‐negative and Gram‐positive eubacteria in their lack of a Leu residue at the −3 position. Antibodies to synthetic peptides that were deduced from two fusions to predicted lipoproteins, identified corresponding amphiphilic membrane proteins of 57 kDa and 78 kDa expressed in the mycoplasma. The P57 sequence contained a proline‐rich N‐terminal region analogous to an adhesin ofMycoplasma gallisepticum. The P78 protein was identical to a serologically defined phase‐variant surface lipoprotein. TnphoAmutagenesis provides an efficient means of systematically characterizing functionally diverse lipoproteins and other exporte

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.mmi_18020283.x

出版商:Blackwell Science Ltd    

年代:1995

数据来源: WILEY