Cristobal Alhambra

Hydride transfer catalyzed by xylose isomerase: Mechanism and quantum effects

We have applied molecular dynamics umbrella‐sampling simulation and ensemble‐averaged variational transition state theory with multidimensional tunneling (EA‐VTST/MT) to calculate the reaction rate of xylose‐to‐ xylulose isomerization catalyzed by xylose isomerase in the presence of two Mg2+ ions. The calculations include determination of the free energy of activation profile and ensemble averaging in the transmission coefficient. The potential energy function is approximated by a combined QM/MM/SVB method involving PM3 for the quantum mechanical (QM) subsystem, CHARMM22 and TIP3P for the molecular mechanical (MM) environment, and a simple valence bond (SVB) local function of two bond distances for the hydride transfer reaction. The simulation confirms the essential features of a mechanism postulated on the basis of kinetics and X‐ray data by Whitlow et al. (Whitlow, M.; Howard, A. J.; Finzel, B. C.; Poulos, T. L.; Winborne, E.; Gilliland, G. L. Proteins 1991, 9, 153) and Ringe, Petsko, and coworkers (Labie, A.; Allen, K.‐N.; Petsko, G. A.; Ringe, D. Biochemistry 1994, 33, 5469). This mechanism involves a rate‐determining 1,2‐hydride shift with prior and post proton transfers. Inclusion of quantum mechanical vibrational energy is important for computing the free energy of activation, and quantum mechanical tunneling effects are essential for computing kinetic isotope effects (KIEs). It is found that 85% of the reaction proceeds by tunneling and 15% by overbarrier events. The computed KIE for the ratio of hydride to deuteride transfer is in good agreement with the experimental results. The molecular dynamics simulations reveal that proton and hydride transfer reactions are assisted by breathing motions of the mobile Mg2+ ion in the active site, providing evidence for concerted motion of Mg2+ during the hydride transfer step.

Inclusion of Quantum Mechanical Vibrational Energy in Reactive Potentials of Mean Force

Classical molecular dynamics and Monte Carlo simulations typically exclude quantum effects on the vibrations of reactants and transition states, and this may lead to significant errors in the computed potential of mean force. To correct this deficiency, a simple approximate procedure is proposed for the inclusion of quantum-mechanical vibrational energy in the computation of reactive potentials of mean force in condensed phases. The method is illustrated by a hydrogen atom transfer and a proton transfer reaction in water, in particular, the 1,5-sigmatropic shift in malonaldehyde and the intermolecular proton shift between ammonium ion and ammonia in an encounter complex. In both cases, quantum-mechanical vibrational energy makes significant contributions by reducing the free energy of activation by 2 to 3 kcal/mol. This finding has important implications in developing empirical potential functions for the study of enzyme reactions, and it is essential to quantize vibrational energy in the computed potenti...

Quantum mechanical tunneling in methylamine dehydrogenase

We report a calculation for a trideuteration kinetic isotope effect (KIE) for the proton transfer step in the oxidation of methylamine by the quinoprotein methylamine dehydrogenase (MADH). The potential field includes 11 025 atoms, and the dynamics are based on a quantum mechanical/molecular mechanical (QM/MM) direct dynamics simulation and canonical variational transition state theory with small-curvature multidimensional tunneling contributions. About 1% of the reaction occurs by overbarrier processes, with the rest due to tunneling, and the calculated KIE is reduced to 5.9 when we omit tunneling. This provides the most striking evidence yet for the contribution of tunneling processes to enzymatic reactions at physiological temperatures.

A hybrid semiempirical quantum mechanical and lattice-sum method for electrostatic interactions in fluid simulations

A method is described to incorporate the Ewald lattice-sum method into quantum mechanical calculations in hybrid quantum and molecular mechanical (QM/MM) fluid simulations. The procedure is illustrated through standard free energy perturbation calculations in the context of Monte Carlo simulations. The free energy of hydration of chloride ion was computed using the hybrid QM/MM-Ewald method, and comparison was made to results obtained with standard spherical cutoff. The results indicate that the hybrid QM/MM-Ewald method can be effectively used to include long-range electrostatic interactions in quantum mechanical calculations of condensed media.

Erratum to: “Quantum mechanical tunneling in methylamine dehydrogenase” [Chem. Phys. Lett. 347 (2001) 512–518]

We report a calculation for a trideuteration kinetic isotope effect (KIE) for the proton transfer step in the oxidation of methylamine by the quinoprotein methylamine dehydrogenase (MADH). The potential field includes 11 025 atoms, and the dynamics are based on a quantum mechanical/molecular mechanical (QM/MM) dynamics simulation and ensemble-averaged canonical variational transition state theory with small-curvature multidimensional tunneling contributions. About 1% of the reaction occurs by overbarrier processes, with the rest due to tunneling. We compute a KIE of 18.3, in good accord with experiment (17.2), but the calculated KIE is reduced to 5.9 when we omit tunneling. This provides the most striking evidence yet for the contribution of tunneling processes to enzymatic reactions at physiological temperatures.

Hydrogen-bonding interactions in the active site of a low molecular weight protein-tyrosine phosphatase

Molecular dynamics simulation of the Michaelis complex, phospho‐enzyme intermediate, and the wild‐type and C12S mutant have been carried out to examine hydrogen‐bonding interactions in the active site of the bovine low molecular weight protein‐tyrosine phosphatase (BPTP). It was found that the Sγ atom of the nucleophilic residue Cys‐12 is ideally located at a position opposite from the phenylphosphate dianion for an inline nucleophilic substitution reaction. In addition, electrostatic and hydrogen‐bonding interactions from the backbone amide groups of the phosphate‐binding loop strongly stabilize the thiolate anion, making Cys‐12 ionized in the active site. In the phospho‐enzyme intermediate, three water molecules are found to form strong hydrogen bonds with the phosphate group. In addition, another water molecule can be identified to form bridging hydrogen bonds between the phosphate group and Asp‐129, which may act as the nucleophile in the subsequent phosphate hydrolysis reaction, with Asp‐129 serving as a general base. The structural difference at the active site between the wild‐type and C12S mutant has been examined. It was found that the alkoxide anion is significantly shifted toward one side of the phosphate binding loop, away from the optimal position enjoyed by the thiolate anion of the wild‐type enzyme in an SN2 process. This, coupled with the high pKa value of an alcoholic residue, makes the C12S mutant catalytically inactive. These molecular dynamics simulations provided details of hydrogen bonding interactions in the active site of BPTP, and a structural basis for further studies using combined quantum mechanical and molecular mechanical potential to model the entire dephosphorylation reaction by BPTP.