Genetic characterization of mannose-sensitive hemagglutinin (MSHA)-negative mutants of Vibrio cholerae derived by Tn5 mutagenesis

Claudia C. Häse, Margaret E. Bauer, Richard A. Finkelstein

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

El Tor biotype Vibrio cholerae strains express a cell-associated mannose-sensitive hemagglutinin (MSHA) which is a putative attachment factor. Several MSHA-negative mutants from V. cholerae strain JBK70 were previously generated by Tn5 mutagenesis [Finn et al., Infect. Immun. 55 (1987) 942-946]. The chromosomal DNA regions containing the Tn5 insertions were isolated from eight strains for further analysis. Nucleotide sequencing of the insertional junctions and corresponding clones containing the intact chromosomal region from the parental strain revealed the presence of several contiguous ORFs. Only two ORFs of this region had received insertions, and these showed remarkable homology to genes involved in the general secretory pathway found in several Gram- bacterial species. Proteins corresponding to the observed ORFs were visualized with the T7 promoter/RNA polymerase expression system. Marker exchange mutagenesis was used to insert kanamycin-resistance cassettes and TnphoA insertions into different locations of this region in the chromosome of wild-type V. cholerae strains. The phenotypes of these mutants showed that this DNA region is involved in MSHA production, but is not required for general extracellular protein secretion.

Original languageEnglish (US)
Pages (from-to)17-25
Number of pages9
JournalGene
Volume150
Issue number1
DOIs
StatePublished - Dec 2 1994
Externally publishedYes

Keywords

  • Bacterial adhesion
  • T7 RNA polymerase
  • TnphoA
  • pili
  • protein secretion
  • pullulanase
  • recombinant DNA
  • toxin
  • transposon-insertion mutants

ASJC Scopus subject areas

  • Genetics

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