Molecular cloning and expression in Escherichia coli K-12 of the gene for a hemagglutinin from Vibrio cholerae.

Abstract

Using antiserum to the purified soluble hemagglutinin we isolated an Escherichia coli K-12 clone expressing the gene for a hemagglutinin from Vibrio cholerae 569B. The plasmid present in this clone was designated pPM471. By deletion analysis with both specific restriction endonucleases and Bal 31 nuclease, we localized the gene, to a 0.72-kilobase region of DNA, implying a molecular weight of less than 27,000 for the protein. Analysis in E. coli K-12 minicells of plasmids containing the cloned gene and deletion derivatives of these plasmids identified a protein of 24,000 daltons correlating with hemagglutinating activity. Using the cloned gene as a probe, we demonstrated the presence of homologous DNA in a variety of V. cholerae strains including both biotypes. Furthermore, by screening gene banks in E. coli K-12 of V. cholerae El Tor O17, we isolated several El Tor clones containing this region of DNA and also expressing hemagglutinating activity.