title: Heat Shock Protein 90 Associates with the Per-Arnt-Sim Domain of Heme-free Soluble Guanylate Cyclase: IMplications for Enzyme Maturation.
abstract: Heat shock protein 90 (hsp90) drives heme insertion into the β1 subunit of soluble guanylate cyclase (sGC) β1, which enables it to associate with a partner sGCα1 subunit and mature into a nitric oxide (NO)-responsive active form. We utilized fluorescence polarization measurements and hydrogen-deuterium exchange mass spectrometry to define molecular interactions between the specific human isoforms hsp90β and apo-sGCβ1. hsp90β and its isolated M domain, but not its isolated N and C domains, bind with low micromolar affinity to a heme-free, truncated version of sGCβ1 (sGCβ1(1-359)-H105F). Surprisingly, hsp90β and its M domain bound to the Per-Arnt-Sim (PAS) domain of apo-sGC-β1(1-359), which lies adjacent to its heme-binding (H-NOX) domain. The interaction specifically involved solvent-exposed regions in the hsp90β M domain that are largely distinct from sites utilized by other hsp90 clients. The interaction strongly protected two regions of the sGCβ1 PAS domain and caused local structural relaxation in other regions, including a PAS dimerization interface and a segment in the H-NOX domain. Our results suggest a means by which the hsp90β interaction could prevent apo-sGCβ1 from associating with its partner sGCα1 subunit while enabling structural changes to assist heme insertion into the H-NOX domain. This mechanism would parallel that in other clients like the aryl hydrocarbon receptor and HIF1α, which also interact with hsp90 through their PAS domains to control protein partner and small ligand binding interactions.
authors.affiliation: From the Departments of Pathobiology.
authors.forename: Anindya
authors.initials: A
authors.lastname: Sarkar
authors.affiliation: From the Departments of Pathobiology.
authors.forename: Yue
authors.initials: Y
authors.lastname: Dai
authors.affiliation: From the Departments of Pathobiology.
authors.forename: Mohammad Mahfuzul
authors.initials: MM
authors.lastname: Haque
authors.affiliation: the Department of Chemistry and Biochemistry, University of Maryland at Baltimore County, Baltimore, Maryland 21250, and.
authors.forename: Franziska
authors.initials: F
authors.lastname: Seeger
authors.affiliation: From the Departments of Pathobiology.
authors.forename: Arnab
authors.initials: A
authors.lastname: Ghosh
authors.affiliation: the Department of Chemistry and Biochemistry, University of Maryland at Baltimore County, Baltimore, Maryland 21250, and.
authors.forename: Elsa D
authors.initials: ED
authors.lastname: Garcin
authors.affiliation: the Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721.
authors.forename: William R
authors.initials: WR
authors.lastname: Montfort
authors.affiliation: Cellular and Molecular Medicine, and.
authors.forename: Stanley L
authors.initials: SL
authors.lastname: Hazen
authors.affiliation: Molecular Cardiology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195.
authors.forename: Saurav
authors.initials: S
authors.lastname: Misra
authors.affiliation: From the Departments of Pathobiology, stuehrd@ccf.org.
authors.forename: Dennis J
authors.initials: DJ
authors.lastname: Stuehr
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D041961:Deuterium Exchange Measurement; D006162:Guanylate Cyclase / Q000737:chemistry* / Q000378:metabolism*; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D006418:Heme / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000378:metabolism*; D000071756:Soluble Guanylyl Cyclase
chemical_list: D018841:HSP90 Heat-Shock Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D006418:Heme; D006162:Guanylate Cyclase; D000071756:Soluble Guanylyl Cyclase
keywords: H-NOX domain; PAS domain; cyclic GMP (cGMP); fluorescence anisotropy; heat shock protein 90 (hsp90); heme insertion; hydrogen exchange mass spectrometry; protein-protein interaction; signal transduction; structural modeling