Resolving the complex role of enzyme conformational dynamics in catalytic function

Abstract

Despite growing evidence suggesting the importance of enzyme conformational dynamics (ECD) in catalysis, a consensus on how precisely ECD influences the chemical step and reaction rates is yet to be reached. Here, we characterize ECD in Cyclophilin A, a well-studied peptidyl-prolyl cis-trans isomerase, using normal and accelerated, atomistic molecular dynamics simulations. Kinetics and free energy landscape of the isomerization reaction in solution and enzyme are explored in unconstrained simulations by allowing significantly lower torsional barriers, but in no way compromising the atomistic description of the system or the explicit solvent. We reveal that the reaction dynamics is intricately coupled to enzymatic motions that span multiple timescales and the enzyme modes are selected based on the energy barrier of the chemical step. We show that Kramers' rate theory can be used to present a clear rationale of how ECD affects the reaction dynamics and catalytic rates. The effects of ECD can be incorporated into the effective diffusion coefficient, which we estimate to be about ten times slower in enzyme than in solution. ECD thereby alters the preexponential factor, effectively impeding the rate enhancement. From our analyses, the trend observed for lower torsional barriers can be extrapolated to actual isomerization barriers, allowing successful prediction of the speedup in rates in the presence of CypA, which is in notable agreement with experimental estimates. Our results further reaffirm transition state stabilization as the main effect in enhancing chemical rates and provide a unified view of ECD's role in catalysis from an atomistic perspective.cis-trans isomerization | Cyclophilin A | enzyme catalysis | enzyme dynamics | Kramers' rate theory E nzymes accelerate reaction rates by several orders of magnitude, allowing them to occur at timescales relevant for cellular functions (1). One of the long-standing issues in biochemistry is how enzymes achieve this remarkable speedup. It is commonly accepted that the most dominating effect arises from significant reduction in the free energy barrier compared to the corresponding noncatalyzed reaction in solution. It is also well established that this predominant effect is mainly electrostatic in nature (2, 3), which is more favorable for the transition state than the reactant or the product (1). However, to what degree and how other factors such as desolvation, steric strain, and enzyme dynamics contribute to catalysis remains disputable. Of particular interest is the role of enzyme dynamics in catalysis that has stirred considerable debate (4-11) partly because it has not been clearly defined, leading to a semantic issue. Also, the link between enzyme dynamics and catalysis is difficult to address both experimentally and theoretically. Currently, the implications of enzyme dynamics are from ensemble-and time-averaged experiments, as the temporal behavior of every atom cannot be observed directly. Although standard molecular dynamics (MD) simulations can provide an a...

Conformational Changes and Free Energies in a Proline Isomerase

“…17 This motion is also observed in the free enzyme, where CypA may adopt at least two distinct conformations including a 4 minor state that has a ~6% population. 18 While previous computational studies on CypA have mostly focused on and provided insight into a potential link between protein motions and overall enzyme function, [19][20][21][22][23][24] we here instead aimed to investigate to what extent simulations can describe quantitatively the free energy surface of the free protein in solution.…”

Perspective: Defining and quantifying the role of dynamics in enzyme catalysis

“…70 The authors concluded that the activation barrier controls the catalysis, and that the dynamical effects are actually anticatalytic by a minor factor of 1/10. Note, however, that this analysis might have some problems, since the considerations of the difference between the enzyme and solution have not correctly reflected the electrostatic differences, which would require a QM/MM description.…”

Dynamical network of residue–residue contacts reveals coupled allosteric effects in recognition, catalysis, and mutation

“…This indicated that overall a new set of contacts were formed and broken upon substrate association. We would like to note that PCA performed on contact space reports on features that are distinct from PCA of fluctuations in Cartesian coordinates (16,42). PCA in contact space defines conformational ensemble in terms of contacts formed and broken and thus yields more information of dynamic changes that take place upon substrate binding or catalysis.…”

“…The suppressed catalytic activity in mutants may be a result of an increase in activation energy and not decreased dynamics (14,15). Although enzyme dynamics may not be responsible for catalytic speed up relative to the uncatalyzed reaction, CypA dynamics has been shown to be coupled to catalytic function (16). Allosteric regulation, i.e., modification of binding or catalysis at the active site due to binding of a ligand at a distal nonoverlapping site, is widespread in biochemical signaling (17,18).…”