CLONING AND CHARACTERIZATION OF TDHA, A LOCUS ENCODING A TONB-DEPENDENT HEME RECEPTOR FROM HAEMOPHILUS-DUCREYI

Haemophilus ducreyi is unable to synthesize heme and most acquire it f rom its only known host, humans. We cloned and sequenced a gene encodi ng an outer membrane receptor for heme. It was designated tdhA (for To nB-dependent heme receptor A) since it was related by sequence homolog y to the family of TonB-dependent receptors. TdhA was strikingly simil ar to open reading frame HI0113 from the genome of Haemophilus influen zae Rd and also shared homology with five other heme receptors, includ ing HxuC, HemR, HmuR, ChuA, and ShuA, from gram-negative bacteria. An Escherichia coli hemA tonB mutant strongly expressing H. ducreyi tdhA grew on low levels of heme as a source of heme only when an intact H. ducreyi Ton system plasmid was present, formally demonstrating functio nal TonB dependence. tdhA was expressed poorly in vitro by H. ducreyi and only under conditions of heme limitation. a survey of H. ducreyi r evealed that all tested strains but one synthesized small amounts of T dhA in vitro under heme-limiting conditions. Surprisingly, an isogenic mutant of tdhA as well as its parent, 35000, both required the same h igh levels of heme for growth (50 mu g/ml [77 mu M] on agar medium). T his result, together with previous findings, suggests that in vitro, t he uptake of heme by H. ducreyi is mediated by a TonB- and TdhA-indepe ndent mechanism, possibly diffusion.