摘要:

SummaryRegulation of gene expression in bacteria, as in eukaryotic cells, is often achieved by variation of mRNA levels. Since the steady state levels of mRNA depend on both the rate of synthesis and the rate of decay, both mechanisms are important for gene regulation. After considerable effort undertaken over many years to understand the regulation of transcription, mRNA degradation has recently gained Increasing attention as an important step in the regulation of some bacterial genes, and many investigations have addressed the mechanisms involved in mRNA decay. ThepufmRNA ofRhodobacter capsulatusencoding pigment binding proteins has become a model system to study decay of a polycistronic mRNA and the role of mRNA degradation in gene expression. Individual segments of the polycistronicpufmRNA display extremely different half‐lives. These differences in stability of mRNA segments are involved in the differential expression ofpufencoded genes and consequently contribute to the stoichiometry of light‐harvesting I and reaction centre complexes that results in optimal growth. In addition, control of mRNA stability is involved in the oxygen‐dependent regulation of photosynthesis genes. High oxygen tension results in decreased stability of the reaction‐centre specificpufmRNA segment, most likely by affecting the rate of endonucleolytic cleavage within the reaction centre coding region. The results obtained from studyingpufmRNA degradation inR. capsulatusandEscherichia colisuggest that a specific distribution of decay promoting and decay impeding mRNA elements along the polycistronic mRNA is responsible for the different half‐lives of individualpu

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01663.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryEscherichia coliis adroit in exploiting environmental energy sources to its greatest profit. A key strategy is to channel electron transport from donors to a terminal acceptor(s) so that the voltage drop is maximal. At the level of transcription, the goal is achieved by the interaction of three global regulatory systems, Fnr, NarL/NarX and ArcB/ArcA. In addition, the regulator FhlA is involved in a cascade‐controlled pathway for the formate branch of the pyruvate fermentation pathwa

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01664.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryRhizobium frediistrain USDA257 produces nitrogen‐fixing nodules on primitive soybean cultivars such as Peking but fails to nodulate agronomically improved cultivars such as McCall. Transposonmutant 257DH4 has two new phenotypes: it nodulates McCall, and its ability to do so is sensitive to the presence of parental strain U5DA257, i.e. it is subject to competitive nodulation blocking. We have isolated a cosmid containing DNA that corresponds to the site of transposon insertion in 257DH4 and have localized Tn5 on an 8.0 kbEcoRI fragment. The 5596 bp DNA sequence that surrounds the insertion site contains seven open reading frames. Five of these, designatednolBTU, ORF4, andnolV, are closely spaced and of the same polarity. nolWandnolXare of the opposite polarity. The initiation codon fornolWlies 155bp upstream from that ofnolB, and it is separated fromnolXby281 bp. The predicted NolT and NolW proteins have putative membrane‐spanning regions. TheN‐terminus of the hypothetical NolW protein also has limited homology to NodH ofRhizobium meliloti, but none of the deduced protein sequences has significant homology to known nodulation gene products. Site‐directed mutagenesis with mudll1734 confirms that inactivation ofnolB, nolT, nolU, nolV, nolW, ornolXextends host range for nodulation to McCall soybean. This phenotype could not be genetically dissected from sensitivity to competitive nodulation blocking. Expression ofnolBTUantinolXis induced as much as 30‐fold by flavonoid signal molecules, even though these genes lacknod‐box promoters. Histochemical staining of McCall roots inoculated withnolB–,nolU–, ornolX–lacZfusions verifies that these genes are expressed continuously from preinfection to the stage of the functional nodule. Although anolU–ORF4–nolVclone hybridizes to a single 8.0 kbEcoRI fragment from 10 strains ofR. frediiand broad‐host‐rangeRhizobiumsp. NGR234, hybridizing sequences are not det

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01665.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryWe investigated the role ofepiQin the biosynthesis of the lantibiotic epidermin.epiQwas essential for epidermin production. It was shown that EpiQ controls epidermin production by transcriptionally activating theepiApromoter, used for transcription of most of the epidermin biosynthetic genes. Additional copies ofepiQincreased epidermin production in the epidermin‐producing wild‐type strainStaphylococcus epidermidisTü3298. TheepiApromoter region was characterized by primer extension analysis. Two inverted repeats, putative operator sites for EpiQ binding, are located upstream of the −35 region and one is localized downstream of the‐10 region. Crude protein extracts from S.epidermidisTü3298 andepiQexpressingEscherichia colicells led to gel mobility shifts of a DNA fragment bearing the inverted repeat which is located immediately upstream of the ‐35 region. DNA fragments bearing the other two inverted repeats were not shifted. TheepiQgene product could be detected by overexpression in theE. coliT7 system using antiserum raised against synthetic pep‐tides of EpiQ. Furthermore, EpiQ, like other DNA‐binding proteins, was shown to bind strongly to he

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01666.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummarySite‐specific insertion of gene cassettes into the insert region of integrons has been demonstrated. Insertion was only observed if the integron DNA integrase was expressed in the recipient cell and if the cassette DNA was ligated prior to transformation. The essential ligation products were resistant to treatment with exonuclease III, indicating that they were closed circular molecules. Insertion of cassettes into integron fragments containing either no insert (one recombination site), or one gene cassette (two recombination sites), was demonstrated. In the latter case, insertion occurred predominantly at the core site located 5′ to the resident cassette, which corresponds to the only site available when no insert is present in the recipient. When DNA molecules including two gene cassettes were used, insertion of only one of the gene cassettes was generally observed, suggesting that resolution of the circular molecule to generate two independent circular cassettes occurred more rapidly than insertion into the recipient integ

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01667.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryManganese superoxide dismutase (thesod Agene product) inEscherichia coli, is negatively regulated by two global regulators, ArcA (aerobic respiration control) and Fur (ferric uptake regulation), coupling its expression to aerobic metabolism and the intracellular iron pool. Footprinting analyses were carried out on thesodApromoter region with purified Fur protein and with ArcA protein overproduced in crude extracts. ArcA is able to bindin vitroin the absence of thein vivotriggering signal. The binding occurs in one step and study of contacts within the operator sequence reveals binding on one side of the double helix. The DNA protection extends to a much larger region (about 65 bp) than would be expected for a 27 kDa protein, suggesting polymerization. Both Fur and ArcA footprints encompass the –35 and –10 promoter region and there is considerable overlap of their binding sequences, in agreement within vivoresults suggesting that either regulator alone can blocksodAtranscription. Furthermore, competition experiments show that Fur and ArcA binding to thesodApromoter are mutually exclusive and that ArcA can easily displace Fur, but not vice versa. The biological significance of thisin vitrobehaviour is discus

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01668.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryThe cells of the extreme thermophileThermus thermophilusare surounded by a regular layer (S‐layer) built up by a protein with an apparent molecular mass of 100 kDa (P100). From purified membrane fractions, three different class of two‐dimensional crystals can be obtained by following alternative extractive procedures. One of these crystals, with p6 symmetry, clearly represents the native S‐layer detected by freeze etching on whole cells, while the other two, showing p2 and p3 symmetries respectively, closely resemble aggregates of bacterial porins. We demonstrate here by limited protreolysis and Western blotting the surprising fact that the protein component of the three crystals is the P100 protein. Our biochemical data also show how this protein forms Ca2+‐stabilized trimers in each crystal, which support the structural analysis that points to p3 units as the common structural block in all of them, and again resembles the situation found in bacterial

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01669.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryThe spatially patterned differentiation of hetero‐cysts in the filamentous cyanobacteriumAnabaenarequires a functionalhetRgene. Transcriptional fusions toluxABshow thathetRis transcribed at a low level throughout the filament whenAnabaenais grown with combined nitrogen, and that induction of the gene begins within 2 h following nitrogen deprivation. By 3.5 h, induction is localized to spaced foci. By 6h, there is an overall induction of at least threefold in whole cultures, reflecting at least a 20‐fold increase within spatially separated cells. The induction requires the presence of a functionalhetRgene, indicating thathetRis autoregulatory. Full induction of a heterocyst structural gene,hepA, also requires a functionalhetRlo

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01670.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryNeisseria meningitidisFAM20 has recently been shown to produce two Fe‐regulated proteins (FrpA and FrpC) related to the RTX family of cytotoxins. Here we report the cloning and DNA sequence of the locus containing the gene encoding the larger meningococcal RTX protein FrpC. FrpC was highly similar to FrpA throughout much of the predicted protein, with two main differences. Whereas the FrpA protein had 13 copies of the nine‐amino‐acid repeat units typical of RTX proteins, FrpC had 43 copies. The additional copies in FrpC apparently arose from a threefold tandem amplification of a 600bp DNA fragment encoding the repeats. In addition, thefrpCgene lacked good promoter consensus sequences. An open reading frame (0RF1) of unknown function was found immediately upstream offrpC, suggesting the possibility thatfrpCwas cotranscribed with ORF1. A probable promoter was found 300 bp upstream of ORF1, and it contained a Fur protein‐binding sequence found in the promoters of Fe‐regulatedEscherichia coligenes. DNA upstream of the ORF 1/frpC promoter was homologous to IStO76‐like elements surrounding capsulation loci of strains ofHaemophilus influenzae.A FrpC‐like protein (reactive in immunoblots with monoclonal antibody 9D4; multiple reactive bands of about 200 to 120kDa) was found in five out of eight meningococcal strains but only in one out of 14 otherNeisseria, suggesting that FrpC may participate in the pathogenesis of meningoc

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01671.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY

摘要:

SummaryPS2 is one of two major proteins detected in the culture media of variousCorynebacterium glutamicumstrains. The coding and promoter regions of thecspBgene encoding PS2 were cloned in lambda gt11 using polyclonal antibodies raised against PS2 for screening. Expression of thecspBgene inEscherichia coliled to the production of a major anti‐PS2 labelled peptide of 63 000 Da, corresponding presumably to the mature form of PS2. It was detected in the cytoplasm, periplasm and surrounding medium ofE. coli.Three other slower migrating bands of 65000, 68 000 and 72 000 Da were detected. The largest one probably corresponds to the precursor form of PS2 inE. coli.Analysis of the nucleotide sequence revealed an open reading frame (ORF) of 1533 nucleotides. The deduced 510‐amino‐acid polypeptide had a calculated molecular mass of 55 426 Da. According to the predicted amino acid sequence, PS2 is synthesized with a W‐terminal segment of 30‐amino‐acid residues reminiscent of eukaryotic and prokaryotic signal pep‐tides, and a hydrophobic domain of 21 residues near the C‐terminus. Although no significant homologies were found with other proteins, it appears that some characteristics and the amino acid composition of PS2 share several common features with surface‐layer proteins. ThecspB gene was then disrupted inC. glutamicumby gene replacement. Freeze‐etching electron microscopy performed on the wild‐type strain indicated that the cell wall ofC. glutamicumis covered with an ordered surface of proteins (surface layer, S‐layer) which is in very close contact with other cell‐wall components. These structures are absent from thecspB‐disrupted strain but are present after reintroduction of the cspB gene on a plasmid into this mutant. Thus we demonstrate that the Slayer protein is

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1993.tb01672.x

出版商:Blackwell Publishing Ltd    

年代:1993

数据来源: WILEY