Energetics underlying hemin extraction from human hemoglobin by Staphylococcus aureus

Area of Science:

• Microbiology
• Biochemistry
• Structural Biology

Background:

• Staphylococcus aureus causes severe infections by acquiring iron from hemoglobin (Hb) via the iron-regulated surface-determinant (Isd) system.
• The Isd system utilizes bacterial receptors like IsdH to bind Hb and extract hemin, a crucial iron source.

Purpose of the Study:

• To elucidate the mechanism by which the IsdH^N2N3 tri-domain unit extracts hemin from human hemoglobin.
• To determine the energetic and kinetic parameters governing hemin release from both α- and β-subunits of tetrameric Hb.

Main Methods:

• Utilized a receptor mutant specific for Hb's α-subunit and a stopped-flow transfer assay to study hemin extraction kinetics.
• Employed isothermal titration calorimetry to analyze protein-protein interactions between IsdH^N2N3 and Hb.
• Performed molecular dynamics simulations to investigate receptor-induced structural changes and bond hydration.

Main Results:

• IsdH^N2N3 accelerates hemin release from Hb by up to 13,400-fold at 37 °C, with a specific activation enthalpy.
• Identified two distinct IsdH^N2N3·Hb interfaces, with one dominating binding affinity and contributing significantly to the overall binding free energy.
• Revealed that receptor-induced structural distortions and increased solvation likely facilitate the rate-limiting hydrolytic cleavage of the axial HisF8 Nϵ-Fe³⁺ bond.

Conclusions:

• The IsdH receptor employs a multi-interface mechanism involving structural distortion and enhanced solvation to efficiently extract hemin from hemoglobin.
• This detailed understanding of the hemin extraction process provides insights into bacterial nutrient acquisition strategies and potential therapeutic targets.