摘要:

SummaryTheesglocus is required for the formation of muiti‐cellular fruiting bodies and spores by the developmental bacteriumMyxococcus xanthusStudies have suggested thatesgmutants are defective in the production of an essential signal (E‐signal) used in cell‐cell communication and that E‐signalling is required for the expression of many developmental genes. Recently we have determined that theesglocus encodes components of a branched‐chain keto aciddehydrogenase.amultienzyme complexinvolved in branched‐chain amino acid metabolism in many bacteria and higher organisms. During vegetative growth inM. xanthus.this enzyme complex appears to participate in the production of the branched‐chain fatty acids found in this organism.M. xanthusfatty acids (including the branched‐chain fatty acids) have been observed to have a variety of effects on developing cells. These effects include; (i) the lysis ofM. xanthuscells (autocide activity), (ii) acceleration of the rate of sporulation and (iii) rescue of sporulation by certain development‐defective mutants. These and other results suggest a model in which the branched‐chain fatty acids. Synthesized during growth, are released from cellular phospholipid by a developmentally regulated phospholipase during fruiting‐body formation. This model proposes that one or more of the branched‐chain fatty acids that are released constitutes the E‐signal which must be transmitted between cells to comp

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02290.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryTheesglocus ofMyxococcus xanthusappears to control the production of a signal that must be transmitted between cells for the completion of multicellular development DNA sequence analysis suggested that theesglocus encodes the E1 decarboxylase (composed of E1α and E1β subunits) of a branched‐chain keto acid dehydrogenase (BCKAD) that is involved in branched‐chain amino acid (BCAA) metabolism. The properties of anesg::Tn5 insertion mutant supported this conclusion. These properties include: (i) the growth yield of the mutant was reduced with increasing concentrations of the BCAAs in the medium while the growth yield of wild‐type cells increased, (ii) mutant extracts were deficient in BCKAD activity, and (iii) growth of the mutant in media with short branched‐chain fatty acids related to the expected products of the BCKAD helped to correct the mutant defects in growth, pigmentation and development. TheesgBCKAD appears to be involved in the synthesis of long branched‐chain fatty acids since the mutant contained reduced levels of this class of compounds. Our results are consistent with a model in which theesg‐encoded enzyme is involved in the synthesis of branched‐chain fatty acids during vegetative growth, and these compounds are used later in cell‐cell signalling d

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02291.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryThe membrane topology of theRhizobium meliloti2011 ExoP protein involved in polymerization and export of succinoglycan was analysed by translational fusions oflacZandphoAreporter genes to theexoPgene. Based on this analysis, the ExoP protein could be divided into an N‐terminal domain mainly located in the periplasmic space and a C‐terminal domain located in the cytoplasm. Whereas the C‐terminal domain of ExoP is characterized by a potential nucleotide‐binding motif, the N‐terminal ExoP domain contains the sequence motif‘PX2pX4SPKX11GXMXG1′, which is also present in proteins involved in the determination of O‐antigen chain length.R. melilotistrains carrying mutatedexoP* genes, exclusively encoding the N‐terminal ExoP domain, produced a reduced amount of succinoglycan. This reduction could be suppressed by a mutation in the regulatory geneexoR.The ratio of low‐molecular‐weight to high‐molecular‐weight succinoglycan was significantly increased in theexoP* mutant strain. In theexoP*lexoRmutant strain only low‐molecular‐weight succinoglycan could be detected. Based on sequence homologies and similar hydropathic profiles, the N‐terminal domain of ExoP was proposed to be a member of a protein family thought to be involved in polysaccha

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02292.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryIn the presence of toluene, xylenes and other structural analogues, the regulatory protein XylR, of the family of transcriptional regulators which act in concert with the σ54factor, activate the promoterPuof the TOL (toluene degradation) plasmid pWW0 ofPseudomonas putida.Amino acid changes Val‐219‐Asp and Ala‐220‐Pro, introducing a proline kink at the hinge region between the N‐terminal A domain and the central portion of XylR, resulted in a semi‐constitutive phenotype which mimicked the activating effect of aromatic inducers. This phenotype was further exacerbated by insertingextra amino acid residueswithin the same inter‐domain region. A truncated XylR protein devoid of the signal‐receiving, amino‐terminal portion of the protein stimulated the cognate promoterPuat high levels independently of inducer addition, both inEscherichia coliand inPseudomonas putida.Replacement of the amino‐terminal domain by a heterologous peptide derived from the MS2 virus polymerase resulted in a hybrid protein still able to bind DNA to the same extentin vivoas XylR, but unable to stimulate transcription. These data indicate that a key event in the activation of XyIR by toluene/xylenes is the release of the repression caused by the A domain of the protein on surfaces located at the central dom

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02293.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryStreptococcus pneumoniaeundergoes spontaneous phase variation in colony morphology. Differences in colony opacity have previously been shown to correlate with differences in the ability of organisms to colonize the mucosal surface of the nasopharynx in an animal model. The genetic basis of opacity variation was identified in transformation experiments. A DNA library, from a strain that varies at high frequency, was screened to identify a single clone capable of transforming a transparent recipient strain which varies at low frequency to an opaque phenotype. Analysis of this opacity locus revealed two genes,glpDandglpF, with similarity to genes required for glycerol metabolism in other bacteria. Following the pneumococcalglpF, repetitive intergenic elements, boxes A and C, were identified. These stem‐loop‐forming elements were not present in the same locus of the recipient strain. Although not required for phase variation in colony opacity, the box element was necessary for expression of phase variation at high frequency. Introduction of the box elements during transformation affected colony morphology, possibly by altering expression of a putative regulatory gene downstream from the box element. Mutagenesis within this region confirmed the contribution of the putative regulatory gene to the expression of colony opacity. Growth characteristics of strains generated in this study provide additional evidence for an association of differences in cell wall autolysis and variation in colony opac

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02294.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryLantibiotic and non‐lantibiotic bacteriocins are synthesized as precursor peptides containing N‐terminal extensions (leader peptides) which are cleaved off during maturation. Most non‐lantibiotics and also some lantibiotics have leader peptides of the so‐ called double‐glycine type. These leader peptides share consensus sequences and also a common processing site with two conserved glycine residues In positions ‐1 and 2. The double‐glycine‐type leader peptides are unrelated to the N‐terminal signal sequences which direct proteins across the cytoplasmic membrane via the sec pathway. Their processing sites are also different from typical signal peptidase cleavage sites, suggesting that a different processing enzyme is involved. Peptide bacteriocins are exported across the cytoplasmic membrane by a dedicated ATP‐binding cassette (ABC) transporter. Here we show that the ABC transporter is the maturation protease and that its proteolytic domain resides in the N‐terminal part of the protein. This result demonstrates that the ABC transporter has a dual function: (i) removal of the leader peptide from its substrate, and (ii) translocation of its substrate across the cytoplasmic membrane. This represents a novel strategy for secretion

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02295.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryMyristoylCoA:protein N‐myristoyltransferase (Nmt) catalyses the co‐transiational, covalent attachment of myristate (C14:0) to the amino‐terminal glycine residue of a number of eukaryotic proteins involved in cellular growth and signal transduction. TheNMT1gene is essential for vegetative growth ofSaccharomyces cerevisiae.Studies were carried out to determine if Nmt is also essential for vegetative growth of the pathogenic fungusCandida albicans.A strain ofC. albicanswas constructed in which one copy ofNMTwas partially deleted and disrupted. A Gly‐447 — Asp mutation was Introduced into the secondNMTallele. This mutation produced marked reductions in catalytic efficiency at 24 and 37° C, as judged byin vitrokinetic studies of the wild‐type and mutant enzymes which had been expressed in, and purified from,Escherichia coli.The growth characteristics of isogenicNMT/NMT, NMT/Δnmt, andnmtΔ/nmtG447D C. albicansstrains were assessed under a variety of conditions. Only thenmtδ/nmtG447Dstrain required myristate for growth. This was true at both 24 and 37°C. Palmitate could not substitute for myristate. Incubation ofnmtΔ/nmtG447Dcells at 37° C in the absence of myristate resulted in cell death as observed by the inability to form colonies on media supplemented with 500 μM myristate. Studies in an immunosuppressed‐mouse model ofC. albicansinfection revealed that theNMT/Δnmtstrain produced 100% lethality within 7 d after intravenous administration while the isogenicnmtΔ/nmtG447Gstrain produced no deaths even after 21 d. These observations establish that Nmt is essential for vegetative growth ofC. albicansand suggest that Inhibitors of this acyltransferase may be therapeutically u

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02296.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryThe intracellular bacteriumListeria monocytogenescan invade several types of normally non‐phagocytic cells. Entry into cultured epithelial cells requires the expression ofinIA, the first gene of an operon, comprising two genes:inIA, which encodes internalin, an 800‐amino‐acid protein, andinIB, which encodes a 630‐amino‐acid protein. Several genes homologous toinIAare detected in the genome ofL. monocytogenes; InIBis one of them. We have assessed the role ofinIBIn invasiveness ofL. monocytogenesby constructing isogenic chromosomal deletion mutants in theinIABlocus. Our findings indicate that: i)inIBis required for entry ofL. monocytogenesinto hepatocytes, but not into intestinal epithelial cells; ii)inIBencodes a surface protein; iii) internalin plays a role for entry into some hepatocyte cell lines. These results provide the first insight into the cell tropism displayed byL. mono

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02297.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryWe have previously described studies of a 22 kDa active fragment of the LasA proteinase. In follow‐up studies of LasA, we have discovered the separate existence of a 23 kDa proteinase which shares many of the enzymatic properties of LasA, including the ability to lyse heat‐killed staphylococoi. However, this apparent serine proteinase, which we designate LasD, is distinct from the 22 kDa active LasA protein for the following reasons: (i) the N‐terminal sequence of LasD shares no homology with LasA or the LasA precursor sequence; (ii)Pseudomonas aeruginosaLasA mutant strains AD1825 and FRD2128 do not produce LasA yet produce LasD; and (iii) specific antibodies to each proteinase do not show any cross‐reactivity. LasD appears to be produced as a 30 kDa protein, which is possibly cleaved to produce a 23 kDa active fragment. The purified LasD fragment (23 kDa) shows strong staphylolytic activity only at higher pH conditions, while LasA exhibits staphylolytic activity over a broad pH range, in addition to their ability to cleave at internal diglycine sites, both the LasD and LasA endoproteinases efficiently cleave

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02298.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

SummaryLipopolysaccharides (LPS) of two polymyxin‐resistant (pmr) mutants and the corresponding parent strain ofEscherichia Coliwere chemically analysed for composition and subjected to31P‐NMR (nuclear magnetic resonance) for assessment of phosphate substitution. Whereas the saccharide portions, fatty acids, and phosphate contents were similar in wild‐type and pmr LPS, the latter contained two‐ to threefold higher amounts of 2‐aminoethanol. The pmr LPS also contained 4‐amino‐4‐deoxy‐l‐arabinopyranose (l‐Arap4N), which is normally not a component ofE. coliLPS. This aminopentose has been assigned to be linked to the 4′‐phosphate of lipid A. Comparative31P‐NMR analysis of the de‐O‐acylated LPS of the wild‐type and pmr strains revealed that phosphate groups of the pmr LPS were mainly (71‐79%) diphosphate diesters, which accounted for only 20% in the wild‐type LPS. Diphosphate monoesters were virtually nonexistent in the pmr LPS, whereas they accounted for 42% of all phosphates in wild‐type LPS. In the lipid A of the pmr strains, the 4′‐phosphate was to a significant degree (35%) substituted byl‐Arap4N, whereas in the wild‐type LPS thel‐ArapN was absent. In the pmr lipid A12‐aminoethanol was completely substituting the glycosidic pyrophosphate but not the glycosidic monophosphate, forming a diphosphate diester linkage at this position in 40% of lipid A molecules. In the wild‐type LPS the glycosidic position of lipid A carried mostly unsubstituted monophosphate and pyrophosphate. Thus the polymyxin resistance was shown to be associated, along with the esterification of the lipid A 4′‐monophosphate by aminoarabinose, with extensiv

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1995.tb02299.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY