Identification and Characterization of the Hemophore-Dependent Heme Acquisition System of Yersinia pestis

Abstract

ABSTRACT Yersinia pestis possesses a heme-protein acquisition system (Hmu) that allows it to utilize heme and heme-protein complexes as the sole sources of iron. Analysis of the Y. pestis CO92 genomic sequence revealed a second heme-protein acquisition gene cluster that shares homology with the hemophore-dependent heme acquisition system (Has system) of Serratia marcescens . This locus consisted of the hasR yp receptor gene, the hasA yp hemophore gene, and genes encoding components of the HasA yp dedicated ABC transporter factor ( hasDE yp ), as well as a tonB homologue ( hasB yp ). By using a reconstituted secretion system in Escherichia coli , we showed that HasA yp is a secreted heme-binding protein and that expression of HasA yp is iron regulated in E. coli . The use of a transcriptional reporter fusion showed that the hasRADEB promoter is Fur regulated and has increased activity at 37°C. Hemoglobin utilization via the Has yp system was studied with both E. coli and Y. pestis , for which has and has hmu mutant strains were used. No contribution of the Has system to heme utilization was observed in either E. coli or Y. pestis under the conditions we tested. Previously it was shown that a deletion of the Hmu system had no effect on the virulence of Y. pestis in a mouse model of bubonic plague. An Hmu − Has − double mutant also retained full virulence in this model of infection. This report constitutes the first attempt to investigate the contribution of the hemophore-dependent heme acquisition system in bacterial pathogenicity.