Rate constants and mechanisms of intrinsically disordered proteins binding to structured targets

Calmodulin (CaM) is found to have the capability to bind multiple targets. Investigations on the association mechanism of CaM to its targets are crucial for understanding protein-protein binding and recognition. Here, we developed a structure-based model to explore the binding process between CaM and skMLCK binding peptide. We found the cooperation between nonnative electrostatic interaction and nonnative hydrophobic interaction plays an important role in nonspecific recognition between CaM and its target. We also found that the conserved hydrophobic anchors of skMLCK and binding patches of CaM are crucial for the transition from high affinity to high specificity. Furthermore, this association process involves simultaneously both local conformational change of CaM and global conformational changes of the skMLCK binding peptide. We found a landscape with a mixture of the atypical "induced fit," the atypical "conformational selection," and "simultaneously binding-folding," depending on the synchronization of folding and binding. Finally, we extend our discussions on multispecific binding between CaM and its targets. These association characteristics proposed for CaM and skMLCK can provide insights into multispecific binding of CaM. structure-based model | Calmodulin | mixture binding mechanism | multispecific recognition M any biological processes are driven by protein-protein binding. The large-scale domain rearrangements in proteins have long been recognized to have a critical role in biological function. This flexibility or conformational dynamics also provide a new viewpoint of binding. In addition to the "lock-andkey" binding mechanism, proposed by Fischer to describe the rigid binding in enzyme catalysis (1), two scenarios, considering flexibility during binding, emerged and are referred as "induced fit" and "conformational selection," addressing the critical roles of flexibility in protein recognition (2-6).Calmodulin (CaM) is an ubiquitous Ca 2+ binding protein that is involved in a wide range of cellular Ca 2+ -dependent signaling pathways. With incorporating Ca 2+ ions, Ca 2+ -CaM regulates the activity of many kinds of proteins including protein phosphatase, inositol triphosphate kinase, nitric oxide synthase, protein kinases, nicotinamide adenine dinucleotide kinase, Ca 2+ pumps, and proteins involved in motility (7-9). The binary complex Ca 2+ -CaM is found to have the capability to bind over 300 targets (7)(8)(9). Exploring the molecular mechanism of Ca 2+ -CaM binding to the different targets is crucial for understanding protein-protein multispecific recognition. X-ray crystallography experiments have been performed to resolve Ca 2+ -loaded CaM structure (10). However, complexes of CaM with target enzymes are difficult to study by NMR and the crystallization method, due to the spatial resolution in the experiments. Alternatively, short peptide sequences corresponding to CaM-binding domains are often used to explore CaM-target protein interactions and several studies suggest that these CaM-peptide in...

Section: Binding Process Involves Both the Local And Global Conformatsupporting

confidence: 56%

Molecular mechanism of multispecific recognition of Calmodulin through conformational changes

Liu

Chu

et al. 2017

“…As hinted at by the polarized structure of the TS, it is possible that the N-terminal half of PUMA binds and folds at its subsite, followed by a separate step where the remainder of the helix forms-an example of the dock-andcoalesce model proposed by Zhou et al (55). However, there are Fig.…”

Section: Discussionmentioning

confidence: 96%

Interplay between partner and ligand facilitates the folding and binding of an intrinsically disordered protein

Rogers

Oleinikovas

Shammas

et al. 2014

152

181

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.Protein folding | stopped flow | coarse-grained simulation | protein-protein interactions | BCL-2

“…On the other hand, the presence of ordered conformations can reduce the time for searching the conformational space to reach the final bound structure. From another perspective, the ordered part accounts for the binding affinity, whereas the disordered part is responsible for the specificity (23,75,76). Finally, we further propose that the preferential selection of a partially ordered conformation is a consequence of the balance between kinetics and thermodynamics as well as between affinity and specificity.…”

Section: Discussionmentioning

confidence: 99%

Multiscaled exploration of coupled folding and binding of an intrinsically disordered molecular recognition element in measles virus nucleoprotein

Wang

Chu

Longhi

et al. 2013

105

119

Numerous relatively short regions within intrinsically disordered proteins (IDPs) serve as molecular recognition elements (MoREs). They fold into ordered structures upon binding to their partner molecules. Currently, there is still a lack of in-depth understanding of how coupled binding and folding occurs in MoREs. Here, we quantified the unbound ensembles of the α-MoRE within the intrinsically disordered C-terminal domain of the measles virus nucleoprotein. We developed a multiscaled approach by combining a physics-based and an atomic hybrid model to decipher the mechanism by which the α-MoRE interacts with the X domain of the measles virus phosphoprotein. Our multiscaled approach led to remarkable qualitative and quantitative agreements between the theoretical predictions and experimental results (e.g., chemical shifts). We found that the free α-MoRE rapidly interconverts between multiple discrete partially helical conformations and the unfolded state, in accordance with the experimental observations. We quantified the underlying global folding-binding landscape. This leads to a synergistic mechanism in which the recognition event proceeds via (minor) conformational selection, followed by (major) induced folding. We also provided evidence that the α-MoRE is a compact molten globule-like IDP and behaves as a downhill folder in the induced folding process. We further provided a theoretical explanation for the inherent connections between "downhill folding," "molten globule," and "intrinsic disorder" in IDP-related systems. Particularly, we proposed that binding and unbinding of IDPs proceed in a stepwise way through a "kinetic divide-and-conquer" strategy that confers them high specificity without high affinity. multiscale simulation | hybrid structure-based model | free-energy surface | flexible binding | flexible recognition I ntrinsically disordered proteins (IDPs) are a newly recognized class of naturally abundant, functional proteins that are unable to fold into a well-defined secondary or tertiary structure under physiological conditions of pH and salinity in the absence of a partner (1-3). IDPs perform a wide spectrum of biological functions, often related to molecular recognition, signal transduction, and cell regulation (4). In contrast, canonical proteins with ordered structure mainly carry out their functions in catalysis and membrane transport (5). The functional repertoire of ordered proteins and that of IDPs are complementary to each other (2, 4). Earlier bioinformatics studies predicted that more than 50% of eukaryotic proteins have long disordered regions (1, 6), and the proportion further increases up to 70% in signaling proteins (7). Due to their ubiquitous occurrence in the living world and close relationship with human diseases, IDPs have in recent years become a hot topic in protein science (3,8). One of the most interesting characteristics of IDPs is that they often undergo disorder-to-order transitions upon binding to their partner(s), such as proteins or DNA (9-14), even though some IDP-p...