title: Interplay between partner and ligand facilitates the folding and binding of an intrinsically disordered protein.
abstract: Protein-protein interactions are at the heart of regulatory and signaling processes in the cell. In many interactions, one or both proteins are disordered before association. However, this disorder in the unbound state does not prevent many of these proteins folding to a well-defined, ordered structure in the bound state. Here we examine a typical system, where a small disordered protein (PUMA, p53 upregulated modulator of apoptosis) folds to an α-helix when bound to a groove on the surface of a folded protein (MCL-1, induced myeloid leukemia cell differentiation protein). We follow the association of these proteins using rapid-mixing stopped flow, and examine how the kinetic behavior is perturbed by denaturant and carefully chosen mutations. We demonstrate the utility of methods developed for the study of monomeric protein folding, including β-Tanford values, Leffler α, Φ-value analysis, and coarse-grained simulations, and propose a self-consistent mechanism for binding. Folding of the disordered protein before binding does not appear to be required and few, if any, specific interactions are required to commit to association. The majority of PUMA folding occurs after the transition state, in the presence of MCL-1. We also examine the role of the side chains of folded MCL-1 that make up the binding groove and find that many favor equilibrium binding but, surprisingly, inhibit the association process.
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Joseph M
authors.initials: JM
authors.lastname: Rogers
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Vladimiras
authors.initials: V
authors.lastname: Oleinikovas
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Sarah L
authors.initials: SL
authors.lastname: Shammas
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Chi T
authors.initials: CT
authors.lastname: Wong
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: De Sancho
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Baker
authors.affiliation: Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom jc162@cam.ac.uk.
authors.forename: Jane
authors.initials: J
authors.lastname: Clarke
mesh_terms: D064267:Intrinsically Disordered Proteins / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D056004:Molecular Dynamics Simulation; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D064267:Intrinsically Disordered Proteins; D008024:Ligands; D050505:Mutant Proteins; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D011518:Proto-Oncogene Proteins
keywords: BCL-2; Protein folding; coarse-grained simulation; protein–protein interactions; stopped flow