Characterization of a Vibrio cholerae virulence factor homologous to the family of TonB‐dependent proteins

Abstract

IrgA is an iron‐regulated virulence factor for infection in an animal model with classical Vibrio cholerae strain 0395. We detected gene sequences hybridizing to irgA at high stringency in clinical isolates in addition to 0395, including another classical strain of V. cholerae, three V. cholerae strains of the EI Tor biotype, three non‐O1 isolates of V. cholerae, and individual isolates of Vibrio parahaemolyticus, Vibrio fluvialis, and Vibrio alginolyticus. No hybridization to IrgA was seen with chromosomal DNA from Vibrio vulnificus or Aeromonas hydrophila. To verify that irgA is the structural gene for the major iron‐regulated outer membrane protein of V. cholerae, we determined the amino‐terminal sequence of this protein recovered after gel electrophoresis and demonstrated that it corresponds to the amino acid sequence of IrgA deduced from the nucleotide sequence. Gel electrophoresis showed that two EI Tor strains of V. cholerae had a major iron‐regulated outer membrane protein identical in size and appearance to IrgA in strain 0395, consistent with the findings of DNA hybridization. We have previously suggested that IrgA might be the outer membrane receptor for the V. cholerae siderophore, vibriobactin. Biological data presented here, however, show that a mutation in IrgA had no effect on the transport of vibriobactin and produced no defect in the utilization of iron from ferrichrome, ferric citrate, haemin or haemoglobin. The complete deduced amino acid sequence of IrgA demonstrated homology to the entire class of Escherichia coli TonB‐dependent proteins, particularly Cir. Unlike the situation with Cir, however, we were unable to demonstrate a role for IrgA as a receptor for catechol‐substituted cephalosporins. The role of IrgA in the pathogenesis of V. cholerae infection, its function as an outer membrane receptor, and its potential interaction with a TonB‐like protein in V. cholerae remain to be determined.