title: Conformational Selection in a Protein-Protein Interaction Revealed by Dynamic Pathway Analysis.
abstract: Molecular recognition plays a central role in biology, and protein dynamics has been acknowledged to be important in this process. However, it is highly debated whether conformational changes happen before ligand binding to produce a binding-competent state (conformational selection) or are caused in response to ligand binding (induced fit). Proposals for both mechanisms in protein/protein recognition have been primarily based on structural arguments. However, the distinction between them is a question of the probabilities of going via these two opposing pathways. Here, we present a direct demonstration of exclusive conformational selection in protein/protein recognition by measuring the flux for rhodopsin kinase binding to its regulator recoverin, an important molecular recognition in the vision system. Using nuclear magnetic resonance (NMR) spectroscopy, stopped-flow kinetics, and isothermal titration calorimetry, we show that recoverin populates a minor conformation in solution that exposes a hydrophobic binding pocket responsible for binding rhodopsin kinase. Protein dynamics in free recoverin limits the overall rate of binding.
authors.affiliation: Howard Hughes Medical Institute, Brandeis University, 415 South Street, Waltham, MA 02454; Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454.
authors.forename: Kalyan S
authors.initials: KS
authors.lastname: Chakrabarti
authors.affiliation: Howard Hughes Medical Institute, Brandeis University, 415 South Street, Waltham, MA 02454; Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454.
authors.forename: Roman V
authors.initials: RV
authors.lastname: Agafonov
authors.affiliation: Howard Hughes Medical Institute, Brandeis University, 415 South Street, Waltham, MA 02454; Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454.
authors.forename: Francesco
authors.initials: F
authors.lastname: Pontiggia
authors.affiliation: Howard Hughes Medical Institute, Brandeis University, 415 South Street, Waltham, MA 02454; Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454.
authors.forename: Renee
authors.initials: R
authors.lastname: Otten
authors.affiliation: Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK.
authors.forename: Matthew K
authors.initials: MK
authors.lastname: Higgins
authors.affiliation: Paul Scherrer Institute, 5232 Viligan PSI, Switzerland.
authors.forename: Gebhard F X
authors.initials: GFX
authors.lastname: Schertler
authors.affiliation: Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454. Electronic address: oprian@brandeis.edu.
authors.forename: Daniel D
authors.initials: DD
authors.lastname: Oprian
authors.affiliation: Howard Hughes Medical Institute, Brandeis University, 415 South Street, Waltham, MA 02454; Department of Biochemistry, Brandeis University, 415 South Street, Waltham, MA 02454. Electronic address: dkern@brandeis.edu.
authors.forename: Dorothee
authors.initials: D
authors.lastname: Kern
mesh_terms: D004926:Escherichia coli; D051606:G-Protein-Coupled Receptor Kinase 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D051603:Recoverin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: C496731:RCVRN protein, human; D051603:Recoverin; D051606:G-Protein-Coupled Receptor Kinase 1
keywords: conformational ensemble; conformational selection; energy landscape; molecular recognition dynamics; protein/protein interaction; recoverin