摘要:

SummaryModrfication of proteins at C‐terminal cysteine residue(s) by the isoprenoids farnesyl (C15) and geranylgeranyl (C20) is essential for the biological function of a number of eukaryotic proteins including fungal mating factors and the small, GTP‐binding proteins of the Ras superfamily. Three distinct enzymes, conserved between yeast and mammals, have been identified that prenylate proteins: farnesyl protein transferase, geranylgeranyl protein transferase type I and geranylgeranyl protein transferase type II. Each prenyl protein transferase has its own protein substrate specificity. Much has been learned about the biology, genetics and biochemistry of protein prenylation and prenyl protein transferases through studies of eukaryotic microorganisms, particularlySaccharo‐myces cerevisiae.The functional Importance of protein prenylation was first demonstrated with fungal mating factors. The initial genetic analysis of prenyl protein transferases was inS. cerewisiaewith the isolation and subsequent characterization of mutations in theRAM1, RAM2, CDC43andBET2genes, each of which encodes a prenyl protein transferase subunit. We review here these and other studies on protein prenylation in eukaryotic microbes and how they relate to and have contributed to our knowledge about protein prenylation in all eukaryotic

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00302.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryThe transcription factor sigma‐54 (σ54) is a sequence‐specific DNA‐binding protein that directs RNA polymer a se to a particular class of promoter. The interaction of σ54with promoter DNA has been analysed by protein‐DNA crosslinking and enzymatic and chemical proteolysis. Direct physical evidence for a DNA‐contacting surface within the carboxy‐terminal one‐third of the protein has been obtained. This region of σ54is likely to be close to the surface of the protein, and contacts DNA when either o54 or the σ54‐holoenzyme bind specifically to promoter DNA. The amino‐terminal region of σ54appears to be highly susceptible to proteolysis, and its integrity influences the accessibility towards proteolysis of a second region of σ54, which includes the DNA‐contacting surface. Thus the amino‐terminal region of σ54may have a role in influencing its DNA‐binding properties, the major determinants of which appear to reside in the carboxy‐t

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00303.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryFour mutants ofStaphylococcus aureusstrain Newman that were defective in the fibrinogen receptor (clumping factor) were isolated by transposonTn917mutagenesis. Southern hybridization analysis of the mutants identified transposon‐host DNA junction fragments, one of which was cloned and used to generate a probe to identify and clone the wild‐type clumping factor locus(clfA).The mutants failed to form clumps in soluble fibrinogen and adhered poorly to polymethylmethacrylate (PMMA) coverslips coated with fibrinogen. A single copy of theclfAgene, when introduced into the chromosome of the mutant strains, fuily compiemented the ciumping deficiency of these strains and restored the ability of these mutants to adhere to fibrinogen‐coated PMMA. in addition, the clonedclfAgene on a shuttle plasmid aiiowed the weakiy ciumping strain 8325‐4 to form clumps with the same avidity as the wild‐type strain Newman and also significantly enhanced the adherence of 8325‐4 strains. Thus the formation of clumps in soluble fibrinogen correlated with adherence of bacteria to solid‐phase fibrinogen. TheclfAgene encodes a fibrinogen‐binding protein with an apparent molecular mass ofc.130 kDa. The amino acid sequence of the protein was deduced from the DNA sequence; it was predicted that a 896 residue protein (molecular mass 92 kDa) would be expressed. The putative ClfA protein has features that suggest that it is associated with the ceil surface. Furthermore it contains a novel 308 residue region comprising dipeptide repeats predominantly of Asp and Ser ending 28 residues upstream from the LPXTG motif common to wall‐associated proteins. Significant homology was found between the ClfA protein and the fibronectin‐binding proteins of S.S. aureus, particularly in

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00304.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryThe plasmid‐partition regions of the P1 and P7 plasmid prophages inEscherichia coliare homologues which each encode two partition proteins, ParA and ParB. The equivalent PI and P7 proteins are closely related. In each case, the proteins are encoded by an operon that is autoregulated by the ParA and ParB proteins in concert. This regulation is species‐specific, as the P1 proteins are unable to repress the P7paroperon and vice versa. The homologous ParA proteins are primarily responsible for repression and bind to regions that overlap the operon promoter in both cases. The DNA‐binding domain of the P7 auto‐repressor lies in the amino‐terminal end of the P7 ParA protein. This region includes a helix‐turn‐helix motif that has a clear counterpart in the P1 ParA sequence. However, despite the common regulatory mechanism and the similarity of the proteins involved in repression, the promoter‐operator sequences of these two operons are very different in sequence and organization. The operator is located downstream of the promoter in P1 and upstream of it in P7, and the two regions show little, if any, homology. How these differences may have arisen from a common ancestral fo

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00305.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryA ∼ 56 000 Da membrane glycoprotein purified from epimastigotes ofTrypanosoma cruziwas characterized biochemically and tested for its efficacy to induce protection in mice from a lethal challenge with this protozoan parasite. Immunofluorescence assays with live and formalin‐fixed epimastigotes and trypomastigotes localized the glycoprotein to the flagellum, the body of the parasite, and the cell membrane. Immunoblotting demonstrated the glyco‐protein's presence in nearly equal amounts in all developmental stages of severalT. cruziisolates. Mice immunized with the purified glycoprotein and challenged with 10000 infectious trypomastigote forms of isolate Y survived the controls by up to four days. This significant protection makes this antigen a potential candidate for a multi‐subunit vaccine against7

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00306.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryA gene was cloned fromActinobacillus pleuropneumoniaestrain 4074 by complementation of anaroDstrain ofEscherichia coli.TheE. coligenearoDcodes for a 3‐dehydroquinase enzyme of type I, active in the aromatic biosynthesis pathway. TheA. pleuropneumoniaegene, termedaroQ, displays no base or ami no acid sequence homoiogy toaroDofE. coli.It is instead homologous to the QUTE andqa‐2genes, respectively ofAspergillus niduiansandNeurospora crassa.These genes code for 3‐dehydroquinase enzymes of type ii, involved in the catabolism of quinic acid. The 1.6 kb fragment, which includesaroQ, carries four overlapping or adjacent open reading frames: adapDgene;aroQ;one without homology to sequences in GenBank; and one with homology to the C‐terminal 40% ofchlNof

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00307.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryThe nucleotide sequence of the proximal half of therfbregion ofShigella flexnerihas been determined, and the genes encoding enzymes involved in the biosynthesis of dTDP‐rhamnose have been identified. These genes show strong homology to therfbgenes encoding dTDP‐rhamnose biosynthesis inSalmonella entericaserovartyphimurium(strain LT2) and S.entericaserovaranatum(strain M32) (Jianget al., 1991; Wanget al., 1992). An open reading frame upstream ofrfbBwas also identified which encoded a protein having strong similarity with GalU, and has been designatedgalF.GalF has 92% amino acid sequence identity with anS. entericaLT2 gene,orf2X8, which issimilarly situatedupstream ofrfbB(Jiang et al., 1991). The T7 expression system was utilized to identify proteins corresponding to those predicted from DNA sequence analysis. The similarity of the predicted proteins with proteins that are functionally identical or related, and with others of unknown function from theYersinia enterocoliticaO3rfbregion, and in theEscherichia coliK‐12rffregion are also described. We have re‐addressed the assignment of each gene of the dTDP‐rhamnose pathway with the known enzymes of the pathway, in particularrfbCandrfbD.A reporter plasmid to detect genes encoding enzymes of the dTDP‐rhamnose pathway is described. An analysis of the intergenic region betweengalFandrfbBhas been made, and comparison with the same region fromS. entericaLT

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00308.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryDuring the IF2‐catalysed formation of the 30S initiation complex, the GTP requirement and Its subsequent hydrolysis during 70S complex formation are considered to be essential for translation initiation inEscherichia coli.In order to clarify the role of certain amino acid residues believed to be crucial for the GTP hydrolytic activity ofE. coliIF2, we have introduced seven single amino acid substitutions into its GTP‐binding site (Gly for Val‐400; Thr for Pro‐446; Gly, Glu, Gin for His‐448; and Asn, Glu for Asp‐501). These mutated IF2 proteins were expressedin vivoin physiological quantities and tested for their ability to maintain the growth of anE. colistrain from which the functional chromosomal copy of theinfBgene has been deleted. Only one of the mutated proteins (Asp‐501 to Giu) was able to sustain cell viability and several displayed a dominant negative effect. These results emphasize that the amino acid residues we substituted are essential for the iF2 functions and demonstrate the importance of GTP hydrolysis in translation initiation. These findings are discussed in relation to a previously proposed theoretical model for the

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00309.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryA 15‐17 nucleotide sequence from thegag‐polribosome frameshift site of HIV‐1 directs analogous ribosomal frameshifting inEscherichia coli.Limitation for leucine, which is encoded precisely at the frameshift site, dramatically increased the frequency of leftward frameshifting. Limitation for phenylaianine or arginine, which are encoded just before and just after the frameshift, did not significantly affect frameshifting. Protein sequence analysis demonstrated the occurrence of two closeiy related frameshift mechanisms. In the first, ribosomes appear to bind leucyl‐tRNA at the frameshift site and then slip leftward. This is the 'simultaneous slippage’mechanism. In the second, ribosomes appear to slip before binding amlnoacyl‐tRNA, and then bind phenylaianyl‐tRNA, which is encoded in the left‐shifted reading frame. This mechanism is identicai to the‘overlapping reading’we have demonstrated at other bacterial frameshift sites. The HIV‐1 sequence is prone to frame‐shifting by bo

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00310.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY

摘要:

SummaryRhizobiumspecies elicit the formation of nitrogen‐fixing root nodules through a complex interaction between bacteria and plants. Various bacterial genes involved in the nodulation and nitrogen‐fixation processes have been described and most have been localized on the symbiotic plasmids (pSym). We have found a gene encoding citrate synthase on the pSym plasmid ofRhizobium tropici, a species that forms nitrogen‐fixing nodules on the roots of beans(PhasBoius vuigaris)and trees(Leucaenaspp.). Citrate synthase is a key metabolic enzyme that incorporates carbon into the tricarboxylic acid cycle by catalysing the condensation of acetyl‐CoA and oxalo‐acetic acid to form citrate.R. tropici pcsA(the plasmid citrate synthase gene) is closely related to the corresponding genes of Proteobacteria.pcsAinactivation by aTn5‐mobinsertion causes the bacteria to form fewer nodules (30–50% of the original strain) and to have a decreased citrate synthase activity in minimal medium with sucrose. A clone carrying the pcsA gene complemented ail the phenotypic alterations of thepcsAmutant, and conferredRhizobium iegumino‐sarumbv.phaseoli(which naturally lacks a plasmid citrate synthase gene) a higher nodulation and growth capacity in correlation with a higher citrate synthase activity. We have also found thatpcsAgene expression is sensitive to iron availability, suggesting a possible role ofpcs

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1994.tb00311.x

出版商:Blackwell Publishing Ltd    

年代:1994

数据来源: WILEY