title: Hydrogen-deuterium exchange strategy for delineation of contact sites in protein complexes.
abstract: We use NMR spectra to determine protein-protein contact sites by observing differences in amide proton hydrogen-deuterium exchange in the complex compared to the free protein in solution. Aprotic organic solvents are used to preserve H/D labeling patterns that would be scrambled in water solutions. The binding site between the mammalian co-chaperone Aha1 with the middle domain of the chaperone Hsp90 obtained by our H/D exchange method corresponds well with that in the X-ray crystal structure of the homologous complex from yeast, even to the observation of a secondary binding site. This method can potentially provide data for complexes with unknown structure and for large or dynamic complexes inaccessible via NMR and X-ray methods.
authors.affiliation: Department of Molecular Biology MB2, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, United States. dyson@scripps.edu
authors.forename: H Jane
authors.initials: HJ
authors.lastname: Dyson
authors.forename: Milka
authors.initials: M
authors.lastname: Kostic
authors.forename: Jeff
authors.initials: J
authors.lastname: Liu
authors.forename: Maria A
authors.initials: MA
authors.lastname: Martinez-Yamout
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D018833:Chaperonins; D003903:Deuterium / Q000032:analysis / Q000737:chemistry*; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry*; D006801:Humans; D006859:Hydrogen / Q000032:analysis / Q000737:chemistry*; D018832:Molecular Chaperones / Q000737:chemistry*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D025941:Protein Interaction Mapping / Q000379:methods*; D017434:Protein Structure, Tertiary; D011522:Protons; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry; D012996:Solutions
chemical_list: C475092:AHA1 protein, S cerevisiae; C515652:AHSA1 protein, human; D018841:HSP90 Heat-Shock Proteins; D018832:Molecular Chaperones; D011522:Protons; D029701:Saccharomyces cerevisiae Proteins; D012996:Solutions; D006859:Hydrogen; D003903:Deuterium; D018833:Chaperonins; D004121:Dimethyl Sulfoxide