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A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins.

ABSTRACT: Many intrinsically disordered proteins (IDPs) attain a well-defined structure in a coupled folding and binding reaction with another protein. Such reactions may involve early to late formation of different native structural regions along the reaction pathway. To obtain insights into the transition state for a coupled binding and folding reaction, we performed restrained molecular dynamics simulations using previously determined experimental binding ?b values of the interaction between two IDP domains: the activation domain from the p160 transcriptional co-activator for thyroid hormone and retinoid receptors (ACTR) and the nuclear co-activator binding domain (NCBD) of CREB-binding protein, each forming three well-defined ?-helices upon binding. These simulations revealed that both proteins are largely disordered in the transition state for complex formation, except for two helices, one from each domain, that display a native-like structure. The overall transition state structure was extended and largely dynamic with many weakly populated contacts. To test the transition state model, we combined site-directed mutagenesis with kinetic experiments, yielding results consistent with overall diffuse interactions and formation of native intramolecular interactions in the third NCBD helix during the binding reaction. Our findings support the view that the transition state and, by inference, any encounter complex in coupled binding and folding reactions are structurally heterogeneous and largely independent of specific interactions. Furthermore, experimental ?b values and Brønsted plots suggested that the transition state is globally robust with respect to most mutations but can display more native-like features for some highly destabilizing mutations, possibly because of Hammond behavior or ground-state effects.

SUBMITTER: Karlsson E

PROVIDER: S-EPMC6349112 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins.

Karlsson Elin E Andersson Eva E Dogan Jakob J Gianni Stefano S Jemth Per P Camilloni Carlo C

The Journal of biological chemistry 20181204 4

Many intrinsically disordered proteins (IDPs) attain a well-defined structure in a coupled folding and binding reaction with another protein. Such reactions may involve early to late formation of different native structural regions along the reaction pathway. To obtain insights into the transition state for a coupled binding and folding reaction, we performed restrained molecular dynamics simulations using previously determined experimental binding Φ<sub>b</sub> values of the interaction between ...[more]

PMID: 30514761

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Project description:Successful extensions of protein-folding energy landscape theory to intrinsically disordered proteins' (IDPs') binding-coupled-folding transition can enormously simplify this biomolecular process into diffusion along a limited number of reaction coordinates, and the dynamics subsequently is described by Kramers' rate theory. As the critical pre-factor, the diffusion coefficient D has direct implications on the binding kinetics. Here, we employ a structure-based model (SBM) to calculate D in the binding-folding of an IDP prototype. We identify a strong position-dependent D during binding by applying a reaction coordinate that directly measures the fluctuations in a Cartesian configuration space. Using the malleability of the SBM, we modulate the degree of conformational disorder in an isolated IDP and determine complex effects of intrinsic disorder on D varying for different binding stages. Here, D tends to increase with disorder during initial binding but shows a non-monotonic relationship with disorder in terms of a decrease-followed-by-increase in D during the late binding stage. The salt concentration, which correlates with electrostatic interactions via Debye-Hückel theory in our SBM, also modulates D in a stepwise way. The speeding up of diffusion by electrostatic interactions is observed during the formation of the encounter complex at the beginning of binding, while the last diffusive binding dynamics is hindered by non-native salt bridges. Because D describes the diffusive speed locally, which implicitly reflects the roughness of the energy landscape, we are eventually able to portray the binding energy landscape, including that from IDPs' binding, then to binding with partial folding, and finally to rigid docking, as well as that under different environmental salt concentrations. Our theoretical results provide key mechanistic insights into IDPs' binding-folding, which is internally conformation- and externally salt-controlled with respect to diffusion.

Project description:Gram-positive pathogenic bacteria Staphylococcus express and secret staphylococcal peroxidase inhibitor (SPIN) proteins to help evade neutrophil-mediated immunity by inhibiting the activity of the main oxidative-defense player myeloperoxidase (MPO) enzyme. SPIN contains a structured 3-helix bundle C-terminal domain, which can specifically bind to MPO with high affinity, and an intrinsically disordered N-terminal domain (NTD), which folds into a structured β-hairpin and inserts itself into the active site of MPO for inhibition. Mechanistic insights of the coupled folding and binding process are needed in order to better understand how residual structures and/or conformational flexibility of NTD contribute to the different strengths of inhibition of SPIN homologs. In this work, we applied atomistic molecular dynamics simulations on two SPIN homologs, from S. aureus and S. delphini, respectively, which share high sequence identity and similarity, to explore the possible mechanistic basis for their different inhibition efficacies on human MPO. Direct simulations of the unfolding and unbinding processes at 450 K reveal that these two SPIN/MPO complexes systems follow surprisingly different mechanisms of coupled binding and folding. While coupled binding and folding of SPIN-aureus NTD is highly cooperative, SPIN-delphini NTD appears to mainly utilize a conformational selection-like mechanism. These observations are in contrast to an overwhelming prevalence of induced folding-like mechanisms for intrinsically disordered proteins that fold into helical structures upon binding. Further simulations of unbound SPIN NTDs at room temperature reveal that SPIN-delphini NTD has a much stronger propensity of forming β-hairpin like structures, consistent with its preference to fold and then bind. These may help explain why the inhibition strength is not well correlated with binding affinity for different SPIN homologs. Altogether, our work establishes the relationship between the residual conformational stability of SPIN-NTD and their inhibitory function, which can help us develop new strategies towards treating Staphylococcal infections. Graphical Abstract

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A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins.

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A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins.

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