title: A deconvolution method for the separation of specific versus nonspecific interactions in noncovalent protein-ligand complexes analyzed by ESI-FT-ICR mass spectrometry.
abstract: A method to separate specific and nonspecific noncovalent interactions observed in ESI mass spectra between a protein and its ligands is presented. Assuming noncooperative binding, the specific ligand binding is modeled as a statistical distribution on identical binding sites. For the nonspecific fraction we assume a statistical distribution on a large number of "nonspecific" interacting sites. The model was successfully applied to the noncovalent interaction between the protein creatine kinase (CK) and its ligands adenosine diphosphate (ADP) and adenosine triphosphate (ATP) that both exhibit nonspecific binding in the mass spectrum. The two sequential dissociation constants obtained by applying our method are K(1,diss) = 11.8 +/- 1.5 microM and K(2,diss) = 48 +/- 6 microM for ADP. For ATP, the constants are K(1,diss) = 27 +/- 7 microM and K(2,diss) = 114 +/- 27 microM. All constants are in good correlation with reported literature values. The model should be valuable for systems with a large dissociation constant that require high ligand concentrations and thus have increased potential of forming nonspecific adducts.
authors.affiliation: Laboratoire des Mécanismes Réactionnels, Ecole Polytechnique, Palaiseau, France.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Daubenfeld
authors.forename: Anne-Pascale
authors.initials: AP
authors.lastname: Bouin
authors.forename: Guillaume
authors.initials: G
authors.lastname: van der Rest
mesh_terms: D000244:Adenosine Diphosphate / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry*; D000465:Algorithms; D003198:Computer Simulation; D003402:Creatine Kinase / Q000737:chemistry*; D004789:Enzyme Activation; D008956:Models, Chemical; D025941:Protein Interaction Mapping / Q000379:methods*; D012044:Regression Analysis; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D013379:Substrate Specificity
chemical_list: D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D003402:Creatine Kinase