Abstract
Multiple sequence comparisons of proteins of the LcrD/FlbF family allowed the design of primers that specifically amplify sequences coding for type III secretion components. Amplification of Bordetella pertussis DNA with these primers yielded a fragment that was further used as a probe for screening a genomic library. The nucleotide sequence of a positive clone revealed a 2100-bp gene, called bcrD, which specifies a 75-kDa polypeptide homologous to the Yersinia LcrD protein. Chromosome walking allowed the characterization of a 35-kb DNA segment that contains the entire locus and flanking housekeeping genes. The B. pertussis type III secretion locus consists of more than 30 open reading frames (ORFs), most of which are identical to annotated genes of Bordetella spp and share similarities with known type III secretion genes of related bacteria. In order to assess the function of this locus, we engineered a bcrD null mutant. However, none of the tested phenotypes, such as protein secretion, cellular invasion, cytotoxicity or mouse lung colonization, differentiated the mutant from its parental strain. Studies of bcrD and bscN expressions indicated that, under our experimental conditions, these genes are not expressed in vitro. Rest...Continue Reading
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