Temperature dependence of carbon-13 kinetic isotope effects of importance to global climate change

We report here a theoretical study of the 13C kinetic isotope effect (KIE) and its temperature dependence for the reaction OH + CH4 --> H2O + CH3, the major sink of atmospheric methane in the troposphere. The KIE values at various atmospherically significant temperatures were determined by direct dynamics using variational transition state theory with multidimensional tunneling contributions (VTST/MT). The potential energy surfaces (PESs) were generated by hybrid density functional theory as well as by recently developed doubly hybrid density functional theory methods. Comparisons of our calculated KIEs with experimental data and theoretical values in the literature reveal the critical contributions due to multidimensional tunneling and torsion anharmonicity as well as the critical issue of the choice of internal rotational axis.     

Experimental and theoretical multiple kinetic isotope effects for an SN2 reaction. An attempt to determine transition-state structure and the ability of theoretical methods to predict experimental kinetic isotope effects

The secondary alpha-deuterium, the secondary beta-deuterium, the chlorine leaving-group, the nucleophile secondary nitrogen, the nucleophile (12)C/(13)C carbon, and the (11)C/(14)C alpha-carbon kinetic isotope effects (KIEs) and activation parameters have been measured for the S(N)2 reaction between tetrabutylammonium cyanide and ethyl chloride in DMSO at 30 degrees C. Then, thirty-nine readily available different theoretical methods, both including and excluding solvent, were used to calculate the structure of the transition state, the activation energy, and the kinetic isotope effects for the reaction. A comparison of the experimental and theoretical results by using semiempirical, ab initio, and density functional theory methods has shown that the density functional methods are most successful in calculating the experimental isotope effects. With two exceptions, including solvent in the calculation does not improve the fit with the experimental KIEs. Finally, none of the transition states and force constants obtained from the theoretical methods was able to predict all six of the KIEs found by experiment. Moreover, none of the calculated transition structures, which are all early and loose, agree with the late (product-like) transition-state structure suggested by interpreting the experimental KIEs.     

A new interpretation of chlorine leaving group kinetic isotope effects; a theoretical approach

The chlorine leaving group kinetic isotope effects (KIEs) for the S(N)2 reactions between methyl chloride and a wide range of anionic, neutral, and radical anion nucleophiles were calculated in the gas phase and, in several cases, using a continuum solvent model. In contrast to the expected linear dependence of the chlorine KIEs on the C(alpha)-Cl bond order in the transition state, the KIEs fell in a very small range (1.0056-1.0091), even though the C(alpha)-Cl transition state bond orders varied widely from approximately 0.32 to 0.78, a range from reactant-like to very product-like. This renders chlorine KIEs, and possibly other leaving-group KIEs, less useful for studies of reaction mechanisms than commonly assumed. A partial explanation for this unexpected relationship between the C(alpha)-Cl transition state bond order and the magnitude of the chlorine KIE is presented.     

A theoretical analysis of rate constants and kinetic isotope effects corresponding to different reactant valleys in lactate dehydrogenase

In some enzymatic systems large conformational changes are coupled to the chemical step, in such a way that dispersion of rate constants can be observed in single-molecule experiments, each corresponding to the reaction from a different reactant valley. Under this perspective here we present a computational study of pyruvate to lactate transformation catalyzed by lactate dehydrogenase. The reaction consists of a hydride transfer and a proton transfer that seem to take place concertedly. The degree of asynchronicity and the energy barrier depend on the particular starting reactant valley. In order to estimate rate constants we used a free energy perturbation technique adapted to follow the intrinsic reaction coordinate for several possible reaction paths. Tunneling effects are also obtained with a slightly modified version of the ensemble-averaged variational transition state theory with multidimensional tunneling contributions. According to our results the closure of the active site by means of a flexible loop can lead to the formation of different reactant complexes displaying different features in the disposition of some key residues (such as Arg109), interactions with the substrate and number of water molecules in the active site. The chemical step of the reaction takes place with a different reaction rate from each of these complexes. Finally, primary kinetic isotope effects for replacement of the transferring hydrogen of the cofactor with a deuteride are in good agreement with experimental observations, thus validating our methodology.     

Quantum-instanton evaluation of the kinetic isotope effects

A general quantum-mechanical method for computing kinetic isotope effects is presented. The method is based on the quantum-instanton approximation for the rate constant and on the path-integral Metropolis-Monte Carlo evaluation of the Boltzmann operator matrix elements. It computes the kinetic isotope effect directly, using a thermodynamic integration with respect to the mass of the isotope, thus avoiding the more computationally expensive process of computing the individual rate constants. The method should be more accurate than variational transition-state theories or the semiclassical instanton method since it does not assume a single tunneling path and does not use a semiclassical approximation of the Boltzmann operator. While the general Monte Carlo implementation makes the method accessible to systems with a large number of atoms, we present numerical results for the Eckart barrier and for the collinear and full three-dimensional isotope variants of the hydrogen exchange reaction H + H2 --> H2 + H. In all seven test cases, for temperatures between 250 and 600 K, the error of the quantum instanton approximation for the kinetic isotope effects is less than approximately 10%.     

Experimental and theoretical study of the carbon-13 and deuterium kinetic isotope effects in the Cl and OH reactions of CH3F

A laser flash photolysis-resonance fluorescence technique has been employed to determine absolute rate coefficients for the CH3F + Cl reaction in N2 bath gas in the temperature range of 200-700 K and pressure range of 33-133 hPa. The data were fitted to a modified Arrhenius expression k(T) = 1.14 x 10(-12) x (T/298)2.26 exp{-313/T}. The OH and Cl reaction rates of (13)CH3F and CD3F have been measured by long-path FTIR spectroscopy relative to CH3F at 298 +/- 2 K and 1013 +/- 10 hPa in purified air. The FTIR spectra were fitted using a nonlinear least-squares spectral fitting method including line data from the HITRAN database and measured infrared spectra as references. The relative reaction rates defined by alpha = k(light)/k(heavy) were determined to be k(OH+CH3F)/k(OH+CD3F) = 4.067 +/- 0.018, k(OH+CH3F)/k(OH+(13)CH3F) = 1.067 +/- 0.006, k(Cl+CH3F)/k(Cl+CD3F) = 5.11 +/- 0.07, and k(Cl+CH3F)/k(Cl+(13)CH3F) = 1.016 +/- 0.006. The carbon-13 and deuterium kinetic isotope effects in the OH and Cl reactions of CH3F have been further investigated by quantum chemistry methods and variational transition state theory.     

Chlorine kinetic isotope effects on the haloalkane dehalogenase reaction.

We have found chlorine kinetic isotope effects on the dehalogenation catalyzed by haloalkane dehalogenase from Xanthobacter autotrophicus GJ10 to be 1.0045 +/- 0.0004 for 1,2-dichloroethane and 1.0066 +/- 0.0004 for 1-chlorobutane. The latter isotope effect approaches the intrinsic chlorine kinetic isotope effect for the dehalogenation step. The intrinsic isotope effect has been modeled using semiempirical and DFT theory levels using the ONIOM QM/QM scheme. Our results indicate that the dehalogenation step is reversible; the overall irreversibility of the enzyme-catalyzed reaction is brought about by a step following the dehalogenation.     

Proton-coupled electron transfer in soybean lipoxygenase: dynamical behavior and temperature dependence of kinetic isotope effects

The dynamical behavior and the temperature dependence of the kinetic isotope effects (KIEs) are examined for the proton-coupled electron transfer reaction catalyzed by the enzyme soybean lipoxygenase. The calculations are based on a vibronically nonadiabatic formulation that includes the quantum mechanical effects of the active electrons and the transferring proton, as well as the motions of all atoms in the complete solvated enzyme system. The rate constant is represented by the time integral of a probability flux correlation function that depends on the vibronic coupling and on time correlation functions of the energy gap and the proton donor-acceptor mode, which can be calculated from classical molecular dynamics simulations of the entire system. The dynamical behavior of the probability flux correlation function is dominated by the equilibrium protein and solvent motions and is not significantly influenced by the proton donor-acceptor motion. The magnitude of the overall rate is strongly influenced by the proton donor-acceptor frequency, the vibronic coupling, and the protein/solvent reorganization energy. The calculations reproduce the experimentally observed magnitude and temperature dependence of the KIE for the soybean lipoxygenase reaction without fitting any parameters directly to the experimental kinetic data. The temperature dependence of the KIE is determined predominantly by the proton donor-acceptor frequency and the distance dependence of the vibronic couplings for hydrogen and deuterium. The ratio of the overlaps of the hydrogen and deuterium vibrational wavefunctions strongly impacts the magnitude of the KIE but does not significantly influence its temperature dependence. For this enzyme reaction, the large magnitude of the KIE arises mainly from the dominance of tunneling between the ground vibronic states and the relatively large ratio of the overlaps between the corresponding hydrogen and deuterium vibrational wavefunctions. The weak temperature dependence of the KIE is due in part to the dominance of the local component of the proton donor-acceptor motion.     

Linear free energy relationship and kinetic isotope effects as measures for the transition state variation. A computational study

Linear free energy relationship (LFER) and kinetic isotope effects (KIEs) are frequently used experimental means to study reaction mechanisms, in particular the nature of transition states (TSs). Density functional theory (B3LYP/6-311+G**) calculations were carried out on a model reaction, acid-catalyzed ionization of phenylethyl alcohol, to analyze how experimentally observable properties, such as nonlinearity in the Hammett and Br?nsted relations and variation in KIE, are related to a variation of the transition state structure and the mechanism. Several conclusions and insights were obtained: (1) Linear Hammett plots with a dual parameter treatment may not be evidence for an invariable TS structure for a series of reactions. (2) Variations of KIEs indeed reflect the variations of TS structures. (3) Nonlinear Br?nsted plots cannot always be taken as evidence for a stepwise mechanism. (4) A TS structure in the gas phase may change much more easily than a TS structure in solution.     

Computational study of the effects of protein tyrosine nitrations on the catalytic activity of human thymidylate synthase

Tyrosine nitration is a widespread post-translational modification capable of affecting both the function and structure of the host protein molecule. Enzyme thymidylate synthase (TS), a homodimer, is a molecular target for anticancer therapy. Recently purified TS preparations, isolated from mammalian tissues, were found to be nitrated, suggesting this modification to appear endogenously in normal and tumor tissues. Moreover, human TS (hTS) nitration in vitro led to a by twofold lowered catalytic activity following nitration in average of 1 tyrosine residue per monomer (D?browska-Ma? et al. in Org Biomol Chem 10:323–331, 2012), with the modification identified by mass spectrometry at seven different sites (Y33, Y65, Y135, Y213, Y230, Y258 and Y301). In the present paper, combined computational approach, including molecular and essential dynamics and free energy computations, was used to predict the influence on the activity of hTS of nitration of each of the seven tyrosine residues. The simulations were based on the crystal structure of hTS ternary complex with dUMP and Tomudex (PDB code: 1I00), with the Tomudex molecule replaced by the molecule of TS cofactor analogue, tetrahydrofolate. The present results indicate that while with nitration of five out of seven residues (Y33, Y135, Y230, Y258 and Y301), single residue modification appears to have a strong reducing effect on the activity, with the remaining two, Y65 and Y213, no or a weaker influence is apparent. Taken together, these results demonstrate that tyrosine nitrations in the hTS enzyme show clear tendency to influence the structure and dynamics and, in turn, catalytic properties of the host enzyme. These effects are overall distance-dependent.     

A computational study of the effect of bending on secondary kinetic isotope effects in SN2 transition states

Using conventional transition state theory, the secondary deuterium kinetic isotope effect (KIE) in the inversion SN2 reaction of CH3F and F- is calculated to be small, 0.98 (T = 298 K). This is shown to be the result of a balance among opposing entropy and enthalpy terms. By contrast, KIE in the retention SN2 mechanism is calculated to be large (1.5). Accordingly, KIE is a potential observable for discriminating between the two mechanisms. Large KIE's are also found for the inversion and retention mechanisms of the ion pair reactions between CH3F and LiF. All of the transition structures leading to large KIE's have a bent FCF angle and an imaginary frequency that is sensitive to deuterium labeling.     

Extending limits of chlorine kinetic isotope effects

Chlorine kinetic isotope effects exceeding semiclassical limits were observed in enzyme-catalyzed reactions, but their source has not been yet identified. Herein we show that unusually large chlorine kinetic isotope effects are associated with reactions in which chlorine is the central atom that is being passed between two heavy atoms. The origin of these large values is the ratio of imaginary frequencies for light-to-heavy species (the so-called temperature-independent factor).     

A theoretical investigation of alpha-carbon kinetic isotope effects and their relationship to the transition-state structure of S(N)2 reactions

[reaction: see text] The transition structures and alpha-carbon 12C/13C kinetic isotope effects for 22 S(N)2 reactions between methyl chloride and a wide variety of nucleophiles have been calculated using the B1LYP/aug-cc-pVDZ level of theory. Anionic, neutral, and radical anion nucleophiles were used to give a wide range of S(N)2 transition states so the relationship between the magnitude of the alpha-carbon kinetic isotope effect and transition-state structure could be determined. The results suggest that the alpha-carbon 12C/13C kinetic isotope effects for S(N)2 reactions will be large (near the experimental maximum) and that the curve relating the magnitude of the KIE to the percent transfer of the alpha-carbon from the nucleophile to the leaving group in the transition state has a broad maximum. This means very similar KIEs will be found for early, symmetric, and late transition states and that one cannot use the magnitude of these KIEs to estimate transition-state structure.     

The transition state in the reduction of ketone by lithium tri-t-butoxyaluminohydride. A kinetic isotope effect study.

Analysis of experimental and calculated kinetic isotope effects in the title reaction (LiAl(OBu^t)₃H and LiAl(OBu^t)₃D) leads to the conclusion that the transition state occurs close to the mid-point of the reaction coordinate.     

Carbon-14 kinetic isotope effects in the debromination of dibromocinnamic acid

The kinetic isotope effect (KIE) method was applied to study the mechanism of elimination of bromine from erythro-a,b-dibromocinamic acid. The large ¹⁴C KIE for both a- and b-position of side chain of erythro-a,b-dibromocinamic acid proves that elimination of bromine leading to formation of (E)-cinnamic acid proceeds via E2 mechanism.     

Carbon-14 kinetic isotope effects in the debromination of p-methyldibromocinnamic acid

The kinetic isotope effect (KIE) was applied in the study of the mechanism of bromine elimination from p-methyl-erythro-a,b-dibromocinnamic acid, successively labeled at the a and b carbons. The large ¹⁴C KIE for the b-position and small KIE for the a one of side chain of p-methyl-erythro-a,b-dibromocinamic acid proves that elimination of bromine leading to the formation of (E)-p-methylcinnamic acid proceeds via the E1 mechanism.     

Large concentration effects on the magnitude of secondary alpha-deuterium kinetic isotope effects

Because the secondary alpha-deuterium kinetic isotope effects in some S_N2 and E2 reactions are strongly concentration dependent, isotope effects measured at a single concentration could lead to erroneous conclusions about the mechanisms and transition state structures.     

Analysis of kinetic isotope effects for nonadiabatic reactions

Factors influencing the rates of quantum mechanical particle transfer reactions in many-body systems are discussed. The investigations are carried out on a simple model for a proton transfer reaction that captures generic features seen in more realistic models of condensed phase systems. The model involves a bistable quantum oscillator coupled to a one-dimensional double-well reaction coordinate, which is in turn coupled to a bath of harmonic oscillators. Reactive-flux correlation functions that involve quantum-classical Liouville dynamics for chemical species operators and quantum equilibrium sampling are used to estimate the reaction rates. Approximate analytical expressions for the quantum equilibrium structure are derived. Reaction rates are shown to be influenced significantly by both the quantum equilibrium structure and nonadiabatic dynamics. Nonadiabatic dynamical effects are found to play the major role in determining the magnitude of the kinetic isotope effect for the model transfer reaction.