The IsdC Protein from Staphylococcus aureus Uses a Flexible Binding Pocket to Capture Heme

Abstract

Staphylococcus aureus scavenges heme-iron from host hemoproteins using iron-regulated surface determinant (Isd) proteins. IsdC is the central conduit through which heme is passed across the cell wall and binds this molecule using a NEAr Transporter (NEAT) domain. NMR spectroscopy was used to determine the structure of IsdC in complex with a heme analog, zinc-substituted protoporphyrin IX (ZnPPIX). The backbone coordinates of the ensemble of conformers representing the structure exhibit a root mean square deviation to the mean structure of 0.53 ؎ 0.11 Å . IsdC partially buries protoporphyrin within a large hydrophobic pocket that is located at the end of its ␤-barrel structure. The central metal ion of the analog adopts a pentacoordinate geometry in which a highly conserved tyrosine residue serves as a proximal ligand. Consistent with the structure and its role in heme transfer across the cell wall, we show that IsdC weakly binds heme (K D ‫؍‬ 0.34 ؎ 0.12 M) and that ZnPPIX rapidly dissociates from the protein at a rate of 126 ؎ 30 s ؊1. NMR studies of the apo-form of IsdC reveal that a 3 10 helix within the binding pocket undergoes a flexible to rigid transition as heme is captured. This structural plasticity may increase the efficiency of heme transfer across the cell wall by facilitating protein-protein interactions between apoIsdC and upstream hemoproteins.Staphylococcus aureus is an opportunistic Gram-positive pathogen that causes lethal infections such as toxic shock syndrome, meningitis, and endocarditis (1, 2). The bacterium needs the essential nutrient iron to grow and although the human body contains large quantities of this metal, little is directly available to S. aureus as it is sequestered intracellularly (3) or bound to transferrin and lactoferrin (4, 5). During infections, S. aureus procures iron from heme (protoporphyrin IX ϩ iron), which contains ϳ80% of the total iron in the body (6). Heme-loaded hemoglobin (Hb) 5 is released into the blood plasma by the action of microbial hemolysins that rupture erythrocytes (7). A group of newly discovered proteins called iron-regulated surface determinant (Isd) proteins then scavenge heme and transfer it into the cytoplasm where it is degraded to liberate iron (8, 9). The heme-binding IsdC protein plays an essential role in the transfer of heme across the cell wall peptidylglycan (6, 10). Proteins homologous to IsdC are also present in a number of other important human pathogens (Bacillus anthracis and Listeria monocytogenes) (6, 11). Therefore, compounds that inhibit their ability to capture heme may be useful antibiotics.In Gram-negative bacteria, the process of heme-iron acquisition is reasonably well understood. Heme is captured from hemoproteins or hemophore-hemoprotein complexes by specific outer membrane receptors (5). It is then transferred into the periplasm in a Ton-B-dependent manner, where it is moved across the inner membrane by specific ABC-dependent permeases (5). Heme acquisition mechanisms used by Gram-positive bacteria are only beginning...

The Near-iron Transporter (NEAT) Domains of the Anthrax Hemophore IsdX2 Require a Critical Glutamine to Extract Heme from Methemoglobin

“…The eight ␤-strands of IsdX2 NEAT5 are arranged in two antiparallel ␤-sheets, which form a ␤-sandwich (Fig. 6A), consistent with the immunoglobulin-like fold, as observed in multiple NEAT domain crystal structures from B. anthracis and S. aureus (8,12,16,17,35,45,46).…”

Iron-Regulated Surface Determinant (Isd) Proteins of Staphylococcus lugdunensis

“…S. aureus IsdH NEAT3 (PDB ID 2E7D) and IsdB NEAT2 (PDB ID: 3RTL) were used to model S. lugdunensis IsdB NEAT2 (24,72). The NEAT domain of IsdC was modeled on its orthologue from S. aureus (PDB ID 2O6P) (59,69).…”

The Chlorite Dismutase (HemQ) from Staphylococcus aureus Has a Redox-sensitive Heme and Is Associated with the Small Colony Variant Phenotype

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