Here we present a modified version of the on-the-fly string method for the localization of the minimum free energy path in a space of arbitrary collective variables. In the proposed approach the shape of the biasing potential is controlled by only two force constants, defining the width of the potential along the string and orthogonal to it. The force constants and the distribution of the string nodes are optimized during the simulation, improving the convergence. The optimized parameters can be used for umbrella sampling with a path CV along the converged string as the reaction coordinate. We test the new method with three fundamentally different processes: chloride attack to chloromethane in bulk water, alanine dipeptide isomerization, and the enzymatic conversion of isochorismate to piruvate. In each case the same set of parameters resulted in a rapidly converging simulation and a precise estimation of the potential of mean force. Therefore, the default settings can be used for a wide range of processes, making the method essentially parameter free and more user-friendly.

We present a combination of the string method and a path collective variable for the exploration of the free energy surface associated to a chemical reaction in condensed environments. The on-the-fly string method is employed to find the minimum free energy paths on a multidimensional free energy surface defined in terms of interatomic distances, which is a convenient selection to study bond forming/breaking processes. Once the paths have been determined, a reaction coordinate is defined as a measure of the advance of the system along these paths. This reaction coordinate can be then used to trace the reaction Potential of Mean Force from which the activation free energy can be obtained. This combination of methodologies has been here applied to the study, by means of Quantum Mechanics/Molecular Mechanics simulations, of the reaction catalyzed by guanidinoacetate methyltransferase. This enzyme catalyzes the methylation of guanidinoacetate by S-adenosyl-L-methionine, a reaction that involves a methyl transfer and a proton transfer and for which different reaction mechanisms have been proposed.

M.HhaI is a DNA Methyltransferase from Haemophilus haemolyticus that catalyzes the transfer of a methyl group from S-adenosyl-L-methionine (SAM) to the C 5 position of a cytosine. This enzyme is a paradigmatic model for C 5 DNA Methyltransferases due to its major homology to mammalian enzymes and to the availability of high resolution structures of the DNA-enzyme complex. In spite of the number of experimental and theoretical analysis carried out for this system many mechanistic details remain unraveled. We have used full atomistic classical Molecular Dynamics simulations to explore the protein-SAM-DNA ternary complex where the target cytosine base is flipped out into the active site for both the wild type and Glu119Gln mutant. The relaxed structure was used for a combined Quantum Mechanics/Molecular Mechanics exploration of the reaction mechanism using the string method. Exploration of multidimensional Free Energy Surfaces allowed determining a complete picture of the reaction mechanism in agreement with experimental observations. In our proposal

Registro de acceso restringido Este recurso no está disponible en acceso abierto por política de la editorial. No obstante, se puede acceder al texto completo desde la Universitat Jaume I o si el usuario cuenta con suscripción. Registre d'accés restringit Aquest recurs no està disponible en accés obert per política de l'editorial. No obstant això, es pot accedir al text complet des de la Universitat Jaume I o si l'usuari compta amb subscripció. Restricted access item This item isn't open access because of publisher's policy. The full--text version is only available from Jaume I University or if the user has a running suscription to the publisher's contents.

Enzymatic reactions are complex chemical processes taking place in complex dynamic environments. Theoretical characterization of these reactions requires the determination of the reaction coordinate and the transition state ensemble. This is not an easy task because many degrees of freedom may be involved in principle. We present recent efforts to find good enzymatic reaction coordinates and the implications of these findings in the interpretation of enzymatic efficiency. In particular, we analyze different strategies based on the use of minimum free energy paths and direct localization of the dividing surface on multidimensional free energy surfaces. Another strategy is based on the generation of reactive trajectories, using the transition path sampling method, from which transition state configurations can be harvested. Most of the applications carried out until now coincide to stress the change in the nature of the reaction coordinate, in terms of the participation of the chemical and environmental degrees of freedom, as the reaction advances. The degrees of freedom of the chemical system are dominant at the transition state while environmental participation can be more important at early or late stages of the process.

The elucidation of the catalytic role of LinA dehydrohalogenase in the degradation processes of hexachlorocyclohexane (HCH) isomers is extremely important to further studies on the bioremediation of HCH polluted areas. Herein, QM/MM free energy simulations are employed to provide the details of the dehydrochlorination reaction of two HCH isomers (γ and β). In particular, the role of the protonation state of one of the catalytic residues-His73-is explored. Based on our calculations, two distinct minimum free energy pathways (concerted and stepwise) were found for γ-HCH and β-HCH. The choice of the reaction channel for the dehydrochlorination reactions of γ- and β-HCH was shown to depend on the initial mutual orientations of the reacting species in the active site and the protonation form of His73. The sequential pathway comprises the transfer of the proton (Hδ1) between His73 and Asp25 and subsequently the H1/Cl2 pair elimination from the substrate molecule. Within a concerted mechanism, the dehydrochlorination reaction of γ-/β-HCH is initiated with neutral His73 and the Hδ1 proton is transferred upon final product formation. We found that the concerted pathway for β-HCH results in significantly higher free energy of activation than the stepwise route and therefore can be disregarded as not a feasible mechanism. On the other hand, the reaction that occurs with much lower energetic barrier requires a stronger base (i.e., anionic His73) to abstract the proton (H1) from the substrate molecule. The presence of such transient form of His results in higher energy than the respective Michaelis complex and was observed only in the stepwise pathway for both isomers. Furthermore, we have concluded that both pathways (concerted and stepwise) are feasible for the dehydrochlorination reaction of γ-HCH. The activation free energies obtained from the M05-2X/6-31+G(d,p) corrected path coordinate PMF profiles for the dehydrochlorination reactions of the γ-/β-HCH are in good agreement with the experimental values.

The use of antiviral drugs can promote the appearance of mutations in the target protein that increase the resistance of the virus to the treatment. This is also the case...

SignificanceTransition state theory (TST) is the most popular theory to calculate the rates of enzymatic reactions. However, in some cases TST could fail due to the violation of the nonrecrossing hypothesis at the transition state. In the present work we show that even for one of the most controversial enzymatic reactions—the hydride transfer catalyzed by dihydrofolate reductase—the error associated to TST represents only a minor correction to the reaction rate. Moreover, this error is actually larger for the reaction in solution than in the enzymatic active site. Based on this finding and on previous studies we propose an “enzymatic shielding” hypothesis which encompasses various aspects of the catalytic process.