Identification and characterization of a heme-dedicated periplasmic binding protein in Haemophilus ducreyi

Abstract

In Haemophilus ducreyi, heme uptake likely proceeds via a receptor-mediated process. The initial event involves binding to either of two outer membrane receptors, TdhA and HgbA. Once heme is deposited into the periplasmic space, we hypothesize that a heme-dedicated periplasmic binding protein (hHBP) is responsible for transporting heme across the periplasmic space to the inner membrane. To identify the hHBP, periplasmic extracts were generated from H. ducreyi 35000 grown under high and low heme conditions and subjected to proteome mapping. Peptide sequences of upregulated proteins grown under heme-restrictive conditions were determined by mass spectroscopy. A candidate hHBP was identified as a periplasmic binding protein homologous to YfeA of Yersinia pestis. The gene encoding this protein appears to be in a typical ABC transporter operon. Under iron-limiting conditions, no upregulation of the hHBP expression was observed; however, the purified hHBP was shown to specifically bind heme in a concentration-dependent manner.