摘要:

SummaryThe promiscuous plasmid pLS1 encodes two transacting elements that regulate its copy number: protein RepA and antisense RNA II.In vitrotranscription showed that RNAs for both repressors are synthesized from two promoters,PABandPIIFromPAB, genes encoding RepA (transcriptional repressor) and RepB (initiator of replication) are cotranscribed, the target of RepA being located withinPABMutants inrepAor inPABare still sensitive to RepA. However, cloning of therepAgene in a compatible replicon did not result in incompatibility towards pLS1. FromPII, the 50‐nucleotide RNA II is synthesized. The main incompatibility determinant towards pLS1 corresponds to the coding sequence for RNA II. The RNA II target could be reduced to 21 nucleotides, including the RepB initiation of translation signals. We propose that plasmids of the pLS1 family (pE194, pADB201, and pLB4) share functional and structural characteristics for the regulation of their copy number

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1992.tb00840.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SummaryThe previously uncharacterized third and fourth genes (pulEandpulF) of the pullulanase secretion gene operon ofKlebsiella oxytocastrain UNF5023 are, respectively, predicted to encode a 55 kDa polypeptide with a putative nucleotide‐binding site, and a highly hydrophobic 44 kDa polypeptide that probably spans the cytoplasmic membrane several times. Expression ofpulEin minicells or under the control of a strong bacteriophage T7 promoter resulted in the production of a c. 58 kDa cytopLasmic protein. A representative PulE‐β‐galactosidase hybrid protein created byTnlacmutagenesis was also found mainly in the cytoplasm. These results are in line with the predicted absence from PulE of a region of sufficient hydrophobicity to function as a signal sequence. The PulF polypeptide could not be detected either in minicells or when the gene was transcribed from the T7 promoter, but the acquirement of threepulF‐lacZgene fusions that encoded hybrid proteins with relatively high levels of β‐galactosidase activity indicates that this gene can be transcribed and translated. Gene disruption experiments indicated that bothpulEandpulFare required for pullulanase secretion inEscherichia coliK‐12. Both proteins exhibit considerable homology throughout their entire lengths with other proteins involved in protein secretion, pilin assembly, conjugation and transformation competence in a variety of bacteria. In addition, PulE protein has consensus sequences found in a wide variety of nucleotide‐binding proteins. This study completes the initial characterization of the pullulanase secretion gene operon, which comprises 13 genes that are all essential for the transport of pullulanase across the

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1992.tb00841.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SummaryInsertion sequence IS6120fromMycobacterium smegmatiswas identified by its ability to transpose into different sites in the lambda repressor gene, d857, carried on anEscherichia coli/mycobacteria shuttle plasmid. IS6120is a novel 1.5 kb insertion sequence, which has 24‐bp imperfect terminal inverted repeats and generates 9‐bp duplications of the target DNA following insertion. IS6120is present in at least three copies inM. smegmatisbut was not found in other species, includingMycobacterium tuberculosis.Nucleotide sequence analysis revealed that IS6120contains two open reading frames, one of which encodes a putative transposase with similarities to those found in IS256fromStaphylococcus aureus, IST2fromThiobacillus ferrooxidans, and ISRm3fromRhizobium meliloti.The fact that IS6120does not recognize a consensus target sequence for insertion and has no homologous sequences in the other strains studied makes IS6120useful for transposon mutagenesis in mycobacte

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1992.tb00842.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SummaryEscherichia colihaemolysin (Hlya) is a pore‐forming protein which belongs to the family of ‘Repeat‐toxins’ (RTX)(Loet al., 1989; Lallyet al., 1989; Kraiget al., (1990). A model for the pore‐forming structure of HlyA has been proposed (Ludwiget al., 1991) which consists of eight transmembrane segments all present in this hydrophobic region of HlyA. We report here that two synthetic peptides of 10 and 8 amino acids in length (Pep1 and Pep2, respectively), which are derived from transmembrane segment V, are able to form pores in an artificial lipid bilayer. In addition, Pep1 exhibits strong haemolytic activity when tested on human red blood cells (HRBCs). The haemolytic activity of Pep1 and ofE. colihaemolysin is completely inhibited by antibodies raised aga

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1992.tb00843.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY

摘要:

SummaryIron is an essential nutrient forTrichomonas vaginalisand is acquired via highly specific receptor‐mediated mechanisms from the host. Responses ofT. vaginalisto conditions of iron limitation or iron excess were analysed in order to determine whether iron levels in the growth medium regulate certain properties of the parasite. When compared with organisms grown in excess iron, iron limitation resulted in ≥80% lower rates of protein synthesis and ≥3‐fold decreases in cell densities. These parasites also exhibited generation times of 10 hours, 2.5‐fold longer than organisms grown in the usual complex medium. Iron‐restricted growth also resulted in increased binding of lactoferrin by trichomonads, which paralleled elevated expression of the lactoferrin‐binding receptor protein having a relative molecular mass of 136000 daltons (136 kDa). A Mr126kDa protein was concomitantly repressed in low‐iron‐grown parasites. The greater amounts of lactoferrin bound by iron‐depletedT. vaginalisorganisms corresponded with both the expression of additional receptors onto trichomonal surfaces and increased affinity of the receptor for the lactoferrin molecule. Finally, immunoblot analysis of parasites grown under high‐ and low‐iron conditions using sera from patients with trichomoniasis further revealed the synthesis byT. vaginalisof at least 19 iron‐regulated immunogens, and patients' sera also detected the lactoferrin receptor. These data not only show the overall importance of iron to the biology of this protozoan, but illustrate the in vivo iron modulation of gene expression of the biofunctional lactoferrin rece

ISSN:0950-382X    

DOI:10.1111/j.1365-2958.1992.tb00844.x

出版商:Blackwell Publishing Ltd    

年代:1992

数据来源: WILEY