title: Surface Accessibility and Dynamics of Macromolecular Assemblies Probed by Covalent Labeling Mass Spectrometry and Integrative Modeling.
abstract: Mass spectrometry (MS) has become an indispensable tool for investigating the architectures and dynamics of macromolecular assemblies. Here we show that covalent labeling of solvent accessible residues followed by their MS-based identification yields modeling restraints that allow mapping the location and orientation of subunits within protein assemblies. Together with complementary restraints derived from cross-linking and native MS, we built native-like models of four heterocomplexes with known subunit structures and compared them with available X-ray crystal structures. The results demonstrated that covalent labeling followed by MS markedly increased the predictive power of the integrative modeling strategy enabling more accurate protein assembly models. We applied this strategy to the F-type ATP synthase from spinach chloroplasts (cATPase) providing a structural basis for its function as a nanomotor. By subjecting the models generated by our restraint-based strategy to molecular dynamics (MD) simulations, we revealed the conformational states of the peripheral stalk and assigned flexible regions in the enzyme. Our strategy can readily incorporate complementary chemical labeling strategies and we anticipate that it will be applicable to many other systems providing new insights into the structure and function of protein complexes.
authors.affiliation: Interdisciplinary Research Center HALOmem, Martin Luther University Halle-Wittenberg , Kurt-Mothes-Strasse 3, 06120 Halle/Saale, Germany.
authors.forename: Carla
authors.initials: C
authors.lastname: Schmidt
authors.affiliation: Division of Cell & Molecular Biophysics, King's College London , New Hunt's House, SE1 1UL, London, United Kingdom.
authors.forename: Jamie A
authors.initials: JA
authors.lastname: Macpherson
authors.affiliation: Department of Chemistry, King's College London , 7 Trinity Street, SE1 1DB, London, United Kingdom.
authors.forename: Andy M
authors.initials: AM
authors.lastname: Lau
authors.affiliation: Department of Chemistry, King's College London , 7 Trinity Street, SE1 1DB, London, United Kingdom.
authors.forename: Ken Wei
authors.initials: KW
authors.lastname: Tan
authors.affiliation: Division of Cell & Molecular Biophysics, King's College London , New Hunt's House, SE1 1UL, London, United Kingdom.
authors.forename: Franca
authors.initials: F
authors.lastname: Fraternali
authors.affiliation: Department of Chemistry, King's College London , 7 Trinity Street, SE1 1DB, London, United Kingdom.
authors.forename: Argyris
authors.identifier: 0000-0002-6658-3224
authors.initials: A
authors.lastname: Politis
mesh_terms: D019540:Area Under Curve; D025222:Chloroplast Proton-Translocating ATPases / Q000032:analysis*; D002736:Chloroplasts / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D004047:Diethyl Pyrocarbonate / Q000737:chemistry; D056004:Molecular Dynamics Simulation; D021122:Protein Subunits / Q000032:analysis; D012372:ROC Curve; D018724:Spinacia oleracea / Q000201:enzymology; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D021122:Protein Subunits; D025222:Chloroplast Proton-Translocating ATPases; D004047:Diethyl Pyrocarbonate