Energy landscape views for interplays among folding, binding, and allostery of calmodulin domains

Abstract

Ligand binding modulates the energy landscape of proteins, thus altering their folding and allosteric conformational dynamics. To investigate such interplay, calmodulin has been a model protein.Despite much attention, fully resolved mechanisms of calmodulin folding/binding have not been elucidated. Here, by constructing a computational model that can integrate folding, binding, and allosteric motions, we studied in-depth folding of isolated calmodulin domains coupled with binding of two calcium ions and associated allosteric conformational changes. First, mechanically pulled simulations revealed coexistence of three distinct conformational states: the unfolded, the closed, and the open states, which is in accord with and augments structural understanding of recent single-molecule experiments. Second, near the denaturation temperature, we found the same three conformational states as well as three distinct binding states: zero, one, and two calcium ion bound states, leading to as many as nine states. Third, in terms of the nine-state representation, we found multiroute folding/binding pathways and shifts in their probabilities with the calcium concentration. At a lower calcium concentration, "combined spontaneous folding and induced fit" occurs, whereas at a higher concentration, "binding-induced folding" dominates. Even without calcium binding, we observed that the folding pathway of calmodulin domains can be modulated by the presence of metastable states. Finally, full-length calmodulin also exhibited an intriguing coupling between two domains when applying tension.metal | coarse grained | molecular dynamics | multiscale simulations | force P rotein folding and conformational dynamics have often been characterized by the energy landscape of proteins (1-5). The energy landscape is dependent on the molecular physiochemistry and thus is modulated by many factors, such as chemical modification and ligand binding. Ligand binding, in turn, is dependent on the conformation of proteins. Thus, folding, binding, and allosteric conformational dynamics are mutually correlated. Despite their obvious correlation in concept, it has been very challenging to characterize how they are indeed coupled for any single proteins. Here, we address, in depth, how these three types of dynamics, folding, binding, and allosteric conformational dynamics, are coupled from the energy landscape perspective for a specific protein, calmodulin (CaM).CaM is a ubiquitous calcium-binding messenger protein involved in signal transduction (6) and, more importantly here, has been a model protein to investigate folding, binding, and allostery. Full-length CaM has two nearly symmetric globular domains connected by a flexible central helix (7,8). Each domain is composed of paired EF hands containing two Ca 2+ -binding sites (Fig. 1A). Upon binding to Ca 2+ , each CaM domain undergoes substantial conformational change from a closed state to an open state, exposing a hydrophobic patch that can bind with target proteins and regulate downstream processes (9)....

Molecular mechanism of multispecific recognition of Calmodulin through conformational changes

“…Ca 2+ influences both the inherent flexibility and the conformation of each domain of CaM (SI Appendix, Fig. S9) (43)(44)(45)(46). In our simulations with Ca 2+ , all barrier heights (F1(O-LB), F2(O-LB), F3(O-LB), and F(LB-C)) are decreased (Fig.…”

“…In fact, structural analysis revealed that these hydrophobic residues are buried inside of apo-CaM. Upon Ca 2+ binding, the structure of CaM adopts a more extended dumbbell conformation with the hydrophobic interior exposed to solvent to facilitate the subsequent binding with targets (41)(42)(43). In addition, although Ca 2+ does not interact with the central linker directly, it can control the plasticity of the linker (10).…”

Effects of phosphorylation on the intrinsic propensity of backbone conformations of serine/threonine

“…Such effect of phosphorylation on the intrinsic propensity toward certain local backbone conformations can be especially important for intrinsically disordered proteins, for which the long-range global interactions are less significant compared to those of well-structured proteins. In addition, the intrinsic propensity toward certain local backbone conformations of a certain amino acid is also an important input for some computational models of protein folding/aggregation and structure predictions [11][12][13][14][15][16][17]. For example, recent computational works showed that introducing amino acid-specific parameters to the intrinsic conformational propensity in the all-atom force field can drastically improve protein folding and structure predictions [18,19].…”

Ligand-Induced Changes of the Apparent Transition-State Position in Mechanical Protein Unfolding

“…Biophysical Journal 109(2) 365-372 for ligand-binding proteins (24)(25)(26)(31)(32)(33)(34)(35)(36)(37). Both domains consist of a pair of EF-hand folds, each of which is capable of binding one calcium ion.…”