title: The PRE-Derived NMR Model of the 38.8-kDa Tri-Domain IsdH Protein from Staphylococcus aureus Suggests That It Adaptively Recognizes Human Hemoglobin.
abstract: Staphylococcus aureus is a medically important bacterial pathogen that, during infections, acquires iron from human hemoglobin (Hb). It uses two closely related iron-regulated surface determinant (Isd) proteins to capture and extract the oxidized form of heme (hemin) from Hb, IsdH and IsdB. Both receptors rapidly extract hemin using a conserved tri-domain unit consisting of two NEAT (near iron transporter) domains connected by a helical linker domain. To gain insight into the mechanism of extraction, we used NMR to investigate the structure and dynamics of the 38.8-kDa tri-domain IsdH protein (IsdH(N2N3), A326-D660 with a Y642A mutation that prevents hemin binding). The structure was modeled using long-range paramagnetic relaxation enhancement (PRE) distance restraints, dihedral angle, small-angle X-ray scattering, residual dipolar coupling and inter-domain NOE nuclear Overhauser effect data. The receptor adopts an extended conformation wherein the linker and N3 domains pack against each other via a hydrophobic interface. In contrast, the N2 domain contacts the linker domain via a hydrophilic interface and, based on NMR relaxation data, undergoes inter-domain motions enabling it to reorient with respect to the body of the protein. Ensemble calculations were used to estimate the range of N2 domain positions compatible with the PRE data. A comparison of the Hb-free and Hb-bound forms reveals that Hb binding alters the positioning of the N2 domain. We propose that binding occurs through a combination of conformational selection and induced-fit mechanisms that may promote hemin release from Hb by altering the position of its F helix.
authors: Department of Chemistry and Biochemistry, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; University of California, Los Angeles-Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; Molecular Biology Institute, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA.
authors: Megan
authors: M
authors: Sjodt
authors: Department of Chemistry and Biochemistry, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; University of California, Los Angeles-Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; Molecular Biology Institute, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA.
authors: Ramsay
authors: R
authors: Macdonald
authors: Department of Chemistry and Biochemistry, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA.
authors: Thomas
authors: T
authors: Spirig
authors: Department of Chemistry and Biochemistry, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; University of California, Los Angeles-Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; Molecular Biology Institute, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA.
authors: Albert H
authors: AH
authors: Chan
authors: Menzies Research Institute Tasmania, University of Tasmania, Hobart, Tasmania 7000, Australia.
authors: Claire F
authors: CF
authors: Dickson
authors: Department of Biochemistry and Cell Biology, Rice University, Houston, TX 77251, USA.
authors: Marian
authors: M
authors: Fabian
authors: Department of Biochemistry and Cell Biology, Rice University, Houston, TX 77251, USA.
authors: John S
authors: JS
authors: Olson
authors: Menzies Research Institute Tasmania, University of Tasmania, Hobart, Tasmania 7000, Australia.
authors: David A
authors: DA
authors: Gell
authors: Department of Chemistry and Biochemistry, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; University of California, Los Angeles-Department of Energy Institute of Genomics and Proteomics, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA; Molecular Biology Institute, University of California, Los Angeles, 611 Charles Young Drive East, Los Angeles, CA 90095, USA. Electronic address: rclubb@mbi.ucla.edu.
authors: Robert T
authors: RT
authors: Clubb
mesh_terms: D000942:Antigens, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D000942:Antigens, Bacterial; D006454:Hemoglobins; C512792:IsdH protein, Staphylococcus aureus; D011956:Receptors, Cell Surface
keywords: hemoglobin receptor; iron-regulated surface determinant system; large protein NMR; nuclear magnetic resonance spectroscopy; paramagnetic relaxation enhancement