Reliable studies of enzymatic reactions by combined quantum mechanics /molecular mechanics (QM/MM) approaches, with an ab initio description of the quantum region, presents a major challenge to computational chemists. The main problem is the need for a very large computer time for the evaluation of the QM energy, which in turn makes it extremely challenging to perform proper configurational sampling. A seemingly reasonable alternative is to perform energy minimization studies of the type used in gas phase ab initio studies. However, it is hard to see why such an approach should give reliable results in protein active sites. In order to examine the problems with energy minimization QM/MM approaches we chose the hypothetical reaction of a metaphosphate ion with water in the Ras•GAP complex. This hypothetical reaction served as a simple benchmark reaction. The possible problems with the QM/MM minimization were explored by generating several protein configurations from long MD simulations and using energy minimization and scanning of the reaction coordinates to evaluate the corresponding potential energy surfaces of the reaction for each of these different protein configurations. Comparing these potential energy surfaces, we found major variations of the minima of the different total potential energy surfaces. Furthermore, the reaction energies and activation energies also varied significantly even for similar protein configurations. The specific coordination of a magnesium ion, present in the active center of the protein complex, turned out to influence the energetics of the reaction in a major way and a direct coordination to the reactant leads to an increase of the activation energy by 17 kcal/mol. This study demonstrates that energy minimizations starting from a single protein structure could lead to major errors in calculations of activation free energies and binding free energies. Thus we believe that extensive samplings of the configurational space of the protein are essential for meaningful determination of the energetics of enzymatic reactions. The possible relevance of our conclusion with regard to a recent study of the RasGAP reaction is discussed.

Proton transport (PTR) processes play a major role in bioenergetics and thus it is important to gain a molecular understanding of these processes. At present the detailed description of PTR in proteins is somewhat unclear and it is important to examine different models by using well-defined experimental systems. One of the best benchmarks is provided by carbonic anhydrase III (CA III), because this is one of the few systems where we have a clear molecular knowledge of the rate constant of the PTR process and its variation upon mutations. Furthermore, this system transfers a proton between several water molecules, thus making it highly relevant to a careful examination of the "proton wire" concept. Obtaining a correlation between the structure of this protein and the rate of the PTR process should help to discriminate between alternative models and to give useful clues about PTR processes in other systems. Obviously, obtaining such a correlation requires a correct representation of the "chemistry" of PTR between different donors and acceptors, as well as the ability to evaluate the free energy barriers of charge transfer in proteins, and to simulate long-time kinetic processes. The microscopic empirical valence bond (Warshel, A., and R. M. Weiss. 1980. J. Am. Chem. Soc. 102:6218-6226; and Aqvist, J., and A. Warshel. 1993. Chem. Rev. 93:2523-2544) provides a powerful way for representing the chemistry and evaluating the free energy barriers, but it cannot be used with the currently available computer times in direct simulation of PTR with significant activation barriers. Alternatively, one can reduce the empirical valence bond (EVB) to the modified Marcus' relationship and use semimacroscopic electrostatic calculations plus a master equation to determine the PTR kinetics (Sham, Y., I. Muegge, and A. Warshel. 1999. Proteins. 36:484-500). However, such an approximation does not provide a rigorous multisite kinetic treatment. Here we combine the useful ingredients of both approaches and develop a simplified EVB effective potential that treats explicitly the chain of donors and acceptors while considering implicitly the rest of the protein/solvent system. This approach can be used in Langevin dynamics simulations of long-time PTR processes. The validity of our new simplified approach is demonstrated first by comparing its Langevin dynamics results for a PTR along a chain of water molecules in water to the corresponding molecular dynamics simulations of the fully microscopic EVB model. This study examines dynamics of both models in cases of low activation barriers and the dependence of the rate on the energetics for cases with moderate barriers. The study of the dependence on the activation barrier is next extended to the range of higher barriers, demonstrating a clear correlation between the barrier height and the rate constant. The simplified EVB model is then examined in studies of the PTR in carbonic anhydrase III, where it reproduces the relevant experimental results without the use of any parameter that is specifi...

The nature of proton transduction (PTR) through a file of water molecules, along the gramicidin A (gA) channel, has long been considered as being highly relevant to PTR in biological systems. Previous attempts to model this process implied that the so-called Grotthuss mechanism and the corresponding orientation of the water file plays a major role. The present work reexamines the PTR in gA by combining a fully microscopic empirical valence bond (EVB) model and a recently developed simplified EVB-based model with Langevin dynamics (LD) simulations. The full model is used first to evaluate the free energy profile for a stepwise PTR process. The corresponding results are then used to construct the effective potential of the simplified EVB. This later model is then used in Langevin dynamics simulations, taking into account the correct physics of possible concerted motions and the effect of the solvent reorganization. The simulations reproduce the observed experimental trend and lead to a picture that is quite different from that assumed previously. It is found that the PTR in gA is controlled by the change in solvation energy of the transferred proton along the channel axis. Although the time dependent electrostatic fluctuations of the channel and water dipoles play their usual role in modulating the proton-transfer process (Proc. Natl. Acad. Sci. U.S.A. 1984, 81, 444), the PTR rate is mainly determined by the free energy profile. Furthermore, the energetics of the reorientation of the unprotonated water file do not appear to provide a consistent way of assessing the activation barrier for the PTR process. It seems to us that in the case of gA, and probably other systems with significant electrostatic barriers for the transfer of the proton charge, the PTR rate is controlled by the electrostatic barrier. This finding has clear consequences with regards to PTR processes in biological systems.

Possible molecular implementations of quantum-dot cellular automata (QCA) with mixed-valence ruthenium complexes are discussed. A study of the geometric and electronic structures of three mixed-valence ruthenium dimers has been done using ab initio Hartree-Fock and hybrid density functional methods at the HF/3-21G and B3LYP/3-21G levels of theory. These complexes are representatives of Robin and Day classes I, II, and III. Predicted geometries are compared to experimental data, as well as previous computational studies, where available. The B3LYP method predicts structures in better agreement with experiment than the HF method. The analysis of the orbital energies and localization provides insights into the degree of localization and the Robin-Day classification. They are therefore useful tools for the design of mixed valence compounds for use in molecular QCA.

The light-induced proton transport in bacteriorhodopsin has been considered as a model for other light-induced proton pumps. However, the exact nature of this process is still unclear. For example, it is not entirely clear what the driving force of the initial proton transfer is and, in particular, whether it reflects electrostatic forces or other effects. The present work simulates the primary proton transfer (PT) by a specialized combination of the EVB and the QCFF/PI methods. This combination allows us to obtain sufficient sampling and a quantitative free energy profile for the PT at different protein configurations. The calculated profiles provide new insight about energetics of the primary PT and its coupling to the protein conformational changes. Our finding confirms the tentative analysis of an earlier work (A. Warshel, Conversion of light energy to electrostatic energy in the proton pump of Halobacterium halobium, Photochem. Photobiol. 30 (1979) 285-290) and determines that the overall PT process is driven by the energetics of the charge separation between the Schiff base and its counterion Asp85. Apparently, the light-induced relaxation of the steric energy of the chromophore leads to an increase in the ion-pair distance, and this drives the PT process. Our use of the linear response approximation allows us to estimate the change in the protein conformational energy and provides the first computational description of the coupling between the protein structural changes and the PT process. It is also found that the PT is not driven by twist-modulated changes of the Schiff base's pKa, changes in the hydrogen bond directionality, or other non-electrostatic effects. Overall, based on a consistent use of structural information as the starting point for converging free energy calculations, we conclude that the primary event should be described as a light-induced formation of an unstable ground state, whose relaxation leads to charge separation and to the destabilization of the ion-pair state. This provides the driving force for the subsequent PT steps.