The Staphylococcus Aureus IsdH Receptor Forms a Dynamic Complex with Human Hemoglobin and Triggers Heme Release via Two Distinct Hot Spots

Abstract

Iron is an essential nutrient that is actively acquired by bacterial pathogens during infections. Clinically important Staphylococcus aureus obtains iron by extracting heme from hemoglobin using the IsdH surface receptor; extraction is done by a tri-domain unit that contains its second (N2) and third (N3) NEAT domains, joined by a helical linker. Previous studies found these three domains are necessary for heme transfer and resolved the IsdH:Hb complex, but the underlying mechanism remains unknown. In this work, we probed the mechanism of heme capture by using steered and conventional molecular dynamic simulations to explain NMR and stopped-flow transfer kinetics measurements. An alanine mutagenesis analysis reveals that two subsites trigger heme with the transiently forming receptor-Hb interface by contacting Hb's F-helix. The subsites are located within the N3 and linker domains and appear to play distinct roles stabilizing the heme transfer transition state. Using this information, we present a model of the transfer reaction that provides new insight into the mechanism underpinning microbial heme transfer.