title: Mechanism of pKID/KIX Association Studied by Molecular Dynamics Free Energy Simulations.
abstract: The phosphorylated kinase-inducible domain (pKID) associates with the kinase interacting domain (KIX) via a coupled folding and binding mechanism. The pKID domain is intrinsically disordered when unbound and upon phosphorylation at Ser133 binds to the KIX domain adopting a well-defined kinked two-helix structure. In order to identify putative hot spot residues of binding that could serve as an initial stable anchor, we performed in silico alanine scanning free energy simulations. The simulations indicate that charged residues including the phosphorylated central Ser133 of pKID make significant contributions to binding. However, these are of slightly smaller magnitude compared to several hydrophobic side chains not defining a single dominant binding hot spot. Both continuous molecular dynamics (MD) simulations and free energy analysis demonstrate that phosphorylation significantly stabilizes the central kinked motif around Ser133 of pKID and shifts the conformational equilibrium toward the bound conformation already in the absence of KIX. This result supports a view that pKID/KIX association follows in part a conformational selection process. During a 1.5 μs explicit solvent MD simulation, folding of pKID on the surface of KIX was observed after an initial contact at the bound position of the phosphorylation site was enforced following a sequential process of αA helix association and a stepwise association and folding of the second αB helix compatible with available experimental results.
authors.affiliation: Physik-Department T38, Technische Universität München , James-Franck-Str. 1, 85748 Garching, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Bomblies
authors.affiliation: Physik-Department T38, Technische Universität München , James-Franck-Str. 1, 85748 Garching, Germany.
authors.forename: Manuel P
authors.initials: MP
authors.lastname: Luitz
authors.affiliation: Physik-Department T38, Technische Universität München , James-Franck-Str. 1, 85748 Garching, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Zacharias
mesh_terms: D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D001665:Binding Sites; D050882:CREB-Binding Protein / Q000737:chemistry* / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D009154:Mutation; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D058767:Protein Unfolding; D013816:Thermodynamics
chemical_list: C115163:CREB1 protein, human; D017362:Cyclic AMP Response Element-Binding Protein; D050882:CREB-Binding Protein; C494363:CREBBP protein, human