ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02329.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryTo explain how daughter chromosomes are separated into discrete nucleoids and why chromosomes are partitioned with pole preferences, I propose that differential gene expression occurs during DNA replication inEscherichia coli.This differential gene expression means that the daughter chromosomes have different patterns of gene expression and that cell division is not a simple process of binary fission. Differential gene expression arises from autocatalytic gene expression and creates a separate proteolipid domain around each developing chromosome via the coupled transcription‐translation–insertion of proteins into membranes (transertion). As these domains are immiscible, daughter chromosomes are simultaneously replicated and separated into discrete nucleoids. I also propose that the partitioning relationship between chromosome age and cell age arises because the poles of cells have a proteolipid composition that favours transertion from one nucleoid rather than from the other. This hypothesis forms part of an ensemble of related hypotheses which attempt to explain cell division, differentiation and wall growth in bacteria in terms of the physical properties and interactions of the principal constituents of ce

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02330.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryPore‐forming colicins are a family of protein toxins (Mr40–70kDa) produced byEscherichia coliand related bacteria. They are bactericidal by virtue of their ability to form ion channels in the inner membrane of target cells. They provide a useful means of studying questions such as toxin action, polypeptide translocation across and into membranes, voltage‐gated channels and receptor function. These colicins bind to a receptor in the outer membrane before being translocated across the cell envelope with the aid of helper proteins that belong to nutrient‐uptake systems and the so‐called‘Tol’proteins, the function of which has not yet been properly defined. A distinct domain appears to be associated with each of three steps (receptor binding, translocation and formation of voltage‐gated channels). The Tol‐dependent uptake pathway is described here. The structures and interactions of TolA, B, Q and R have by now been quite clearly defined. Transmembrane α‐helix interactions are required for the functional assembly of theE. coliTol complex, which is preferentially located at contact sites between the inner and outer membranes. The number of colicin translocation sites is about 1000 per cell. The role and the involvement of the OmpF porin (with colicins A and N) have been described in a recent study on the structural and functional interactions of a colicin‐resistant mutant of OmpF. The X‐ray crystal structure of the channel‐forming fragment of colicin A and that of the entire colicin la have provided the basis for biophysical and site‐directed muta‐genesis studies. Thanks to this powerful combination, it has been established that the interaction with the receptor in the outer membrane leads to a very substantial conformational change, as a result of which the N‐terminal domains of colicins interact with the lumen of the OmpF pore and then with the C‐terminal domain of TolA. A molten globular conformation of colicins probably constitutes the intermediate translocation/insertion competent state. Once the pore has formed, the polypeptide chain spans the whole cell envelope. Three distinct steps occur in the last stage of the process: (i) fast binding of the C‐terminal domain to the outer face of the cytoplasmic membrane; (ii) a slow insertion of the polypeptide chain into the outer face of the inner membrane in the absence of Δψ and (iii) a profound reorganization of the helix association, triggered by the transmembrane potential and resulting i

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02331.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryTreponema pallidum, the syphilis spirochaete, has a remarkable ability to evade the humoral and cellular responses it elicits in infected hosts. Although formerly attributed to the presence of an outer coat comprised of serum proteins and/or mucopolysaccharides, current evidence indicates that the immuno‐evasiveness of this bacterium is largely the result of its unusual molecular architecture. Based upon a combination of molecular, biochemical, and ultrastructural data, it is now believed that theT. pallidumouter membrane (OM) contains a paucity of poorly immunogenic transmembrane proteins (‘rare outer membrane proteins’) and that its highly immunogentc proteins are lipoproteins anchored predominantly to the periplasmic leaflet of the cytoplasmic membrane. The presence in theT. pallidumOM of a limited number of transmembrane proteins has profound implications for understanding syphilis pathogenesis as well as treponemal physiology. Two major strategies for molecular characterization of rare outer membrane proteins have evolved. The first involves the identification of candidate OM proteins as fusions withEscherichia colialkaline phosphatase. The second involves the characterization of candidate OM proteins identified in outer membranes isolated from virulentT. pallidum.Criteria to define candidate OM proteins and for definitive identification of rare OM proteins are proposed as a guide for future st

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02332.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryPhage λ, like a number of other large DNA bacterio‐phages and the herpesviruses, produces concatemeric DNA during DNA replication. The concatemeric DNA is processed to produce unit‐length, virion DNA by cutting at specific sites along the concatemer. DNA cutting is coordinated with DNA packaging, the process of translocation of the cut DNA into the preformed capsid precursor, the prohead. A key player in the λ DNA packaging process is the phage‐encoded enzyme terminase, which is involved in (i) recognition of the concatemeric λ DNA; (ii) initiation of packaging, which includes the introduction of staggered nicks atcosNto generate the cohesive ends of virion DNA and the binding of the prohead; (iii) DNA packaging, possibly including the ATP‐driven DNA translocation; and (iv) following translocation, the cutting of the terminalcosNlo complete DNA packaging. To one side ofcosNis the sitecosB, which plays a role in the initiation of packaging; along with ATP,cosBstimulates the efficiency and adds fidelity to the endo‐nuclease activity of terminase in cuttingcosN. cosBis essential for the formation of a post‐cleavage complex with terminase, complex I, that binds the prohead, forming a ternary assembly, complex II. Terminase interacts withcosNthrough its large subunit, gpA, and the small terminase subunit, gpNul, interacts withcosB.Packaging follows complex II formation.cosNis flanked on the other side by the sitecosQ, which is needed for termination, but not initiation, of DNA packaging.cosQis required for cutting of the secondcosN, i.e. thecosNat which termination occurs. DNA packaging in λ has aspects that differ from other λ DNA transactions. Unlike the site‐specific recombination system of λ, for DNA packaging the initial site‐specific protein assemblage gives way to a mobile, translocating complete, and unlike the DNA replication system of λ, the same protein machinery is used for both initiation and translocation

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02333.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryMultiple forms of PilC were found inNeisseria meningitidis(Nm) strains isolated from the oropharynx, blood or cerebrospinal fluid expressing either Class I or Class II pili. PilC expression was observed less frequently in case as opposed to carrier isolates. Moreover, PilC and pili were not always co‐expressed. Several heavily piliated strains had no detectable PilC protein as determined by Western blotting using an antiserum previously used to detect such proteins in adhesive variants (Nassifet al., 1994). Serogroup B strain MC58 produced large numbers of pili, but expressed barely detectable amounts of PilC. A clonal variant of this strain with increased expression of PilC concurrently exhibited increased adherence to Chang conjunctival epithelial cells and human umbilical vein endothelial cells (Huvecs), but with more rapid binding to the former. No alteration in pilin sequence occurred in this variant, suggesting the involvement of PilC in increased adhesion. A Pil‐backswitcher isolated from the hyper‐adherent variant was PilC+but was non‐adherent, indicating that any PilC adherence function requires pilus expression. Parental variant (low PilC) produced pili in bundles that were easily detached from the bacterial surface and were frequently associated with Huvec surfaces after bacteria had been sheared off, but pili infrequently replaced bacteria during infection with the PilC‐expressing variant. The hyper‐adherent variant, which appeared to produce morphologically distinct pilus bundles, was able to withstand considerable shearing force and remained firmly attached to Huvecs. This raises the possibility that the observed hyper‐adherence may arise from better anchorage of pili to the bacterial surface in addition to increased adhesion to some host

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02334.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryACryptococcus neoformansgalactose auxotroph was created by ultraviolet light mutagenesis and complemented with aC. neoformansgenomic library. The translated sequence of the complementing DNA revealed a high degree of simlarity to a number of UDP glucose‐D‐gatactose‐1‐phosphate uridylyitransferases. Expression ofC. neoformans GAL7mRNA followed a pattern similar toSaccharomyces cerevisiaeexpression; it was first observed within 2.5 min of induction and fully induced by 30 min. The gene was completely repressed in the presence of glucose. TheGAL7promoter was isolated and used to construct a promoter cassette. Two genes were tested in this cassette for galactose regulation by creatingGAL7promoter fusions with their coding regions. MFα, which encodes a pheromone, was found to produce filaments only in transformants that were induced by galactose. A second gene, β‐glucuronidase (gusA), which is a commonly used reporter gene, was tested and also found to be expressed. When theGAL7p::GUSfusion was used to quantify inducibitity of theGAL7promoter, the level of enzyme activity was at least 500‐fold greater for cells grown in galactose than for cells grown in glucose. TheGAL7promoter is the first inducible promoter characterized inC neoformansand theGUSgene is the first heterologous gene shown to be expressed in this y

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02335.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryThe transcriptional regulation of theStreptococcus salivarius ptsHandptslgenes coding for the general energy‐coupling proteins HPr and enzyme I of the phosphoenolpyruvate:sugar phosphotransferase system were investigated. These genes form an operon with the gene orderptsH–ptsl.Three distinct mRNA species were detected: a 0.5 kb transcript specific forptsH, and two long transcripts (2.2 arid 2.4 kb) covering the whole pts operon. Transcription of all these mRNAs initiated at the same nucleotide located 9 bp downstream from a promoter located immediately upstream from theptsHgene. The presence of a high‐energy stem–loop structure (T0) located at the begining ofptslwas responsible for the premature transcrrption termination generating the 0.5 kbptsH‐specific transcript. The long transcripts ended in the poly(U) region of two rho‐independent‐like terminators (T1and T2) at the 3′ end ofptsl.Studies with a 2‐deoxyglucose‐resistant spontaneous mutant ofS. salivarius(L26) that produces an HPr–El fusion protein suggest that the regulation of HPr and El expression involves transcriptional as well as tra

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02336.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryPreviously, we have shown that thenodEgene is a major determinant of the difference in host range betweenRhizobium leguminosarumbiovarsviciaeandtrifolii.A new genetic test system for stringent functional analysis ofnodEgenes was constructed. By testing chimericnodEgenes constructed by the exchange of poiymerase chain reaction (PCR)‐generated restriction cassettes, we show that a central domain, containing only 44 non‐conserved amino acid residues, determines the host specificity of the NodE protein (401 amino acid residues). Mass spectrometric analysis of the lipo‐chitin oligosaccharides (LCOs) produced by the new test strain containing the biovarviciae nodEgene shows that molecules containing a polyunsaturated C18:4 (trans‐2.trans‐4.trans‐6.cis‐11‐octadecatetraenoic) fatty acyl moiety are produced, as is the case for wild‐type R.leguminosarumbv.viciae.The LCOs determined by the biovartrifolii nodEgene, which was overproduced in our test strain, carry C1 8:2 and C18:3 fatty acyl chains containing two or three conjugatedtransdouble bonds, respectively. Therefore, the main difference between thenodE‐determined LCOs of biovarviviaeandtrifoliiin this system is the presence or absence of one cis double bond, resulting in the very different hydrophobicity of the LCOs. Using a newly developed spot application assay, we show that the 18:2‐ and C18:3‐containing LCOs are able to induce the formation of nodule primordia on roots ofTrifolium pratense.On the basis of these and other recent results, we propose that the host range of nodulation of theR. leguminosarumbiovarsviciaeandtrifoliiis determined by the degree of hydrophobicity of the poly‐unsaturated fatty acyl moieties of their LCOs, which is mediated by the host‐specific central

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02337.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryThe lincomycin (LM)‐production gene cluster of the overproducing strainStreptomyces iincolnensis78‐11 was cloned, analysed by hybridization, as well as by DNA sequencing, and compared with the respective genome segments of other lincomycin producers. Thelmb/lmrgene cluster is composed of 27 open reading frames with putative biosynthetic or regulatory functions (lmbgenes) and three resistance(lmr)genes, two of which,lmrAandlmrC, flank the cluster. A very similar overall organization of thelmb/lmrcluster seems to be conserved in four other LM producers, although the clusters are embedded in non‐homologous genomic surroundings, in the wild‐type strain (S. lincolnensisNRRL2936), thelmb/lmr‐cluster apparently is present only in single copy. However, in the industrial strainS. lincolnensis78‐11 the non‐adjacent gene clusters for the production of LM and melanin(melC)both are duplicated on a large (0.45‐0.5 Mb) fragment, accompanied by deletion events. This indicates that enhanced gene dosage is one of the factors for the overproduction of LM and demonstrates that large‐scale genome rearrangements can be a result of classical strain improvement by mutagenesis. Only a minority of the putative Lmb proteins belong to known protein families. These include members of the γ‐glutamyl transferases (LmbA), amino acid acylases (LmbC), aromatic amino acid aminotransferases (LmbF), imidazoleglycerolphosphate dehydratases (LmbK), dTDP‐glucose synthases (LmbO), dTDP‐glucose 4,6‐dehydratases (LmbM) and (NDP‐) ketohexose (or ketocyclitol) aminotransferases (LmbS). In contrast to earlier proposals on the biosynthetic pathway of the C‐8 sugar moiety (methylthiolincosaminide), this branch of the LM pathway actually seems to be based on nucleotide

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02338.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY