A Theoretical Analysis of Rate Constants and Kinetic Isotope Effects Corresponding to Different Reactant Valleys in Lactate Dehydrogenase
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- 作者：Silvia Ferrer ; Iñ ; aki Tuñ ; ó ; n ; Sergio Martí ; Vicente Moliner ; Mireia Garcia-Viloca ; À ; ngels Gonzá ; lez-Lafont ; José ; M. Lluch
- 刊名：Journal of the American Chemical Society
- 出版年：2006
- 出版时间：December 27, 2006
- 年：2006
- 卷：128
- 期：51
- 页码：16851 - 16863
- 全文大小：416K
- 年卷期：v.128,no.51(December 27, 2006)
- ISSN：1520-5126
文摘
In some enzymatic systems large conformational changes are coupled to the chemical step, insuch a way that dispersion of rate constants can be observed in single-molecule experiments, eachcorresponding to the reaction from a different reactant valley. Under this perspective here we present acomputational study of pyruvate to lactate transformation catalyzed by lactate dehydrogenase. The reactionconsists of a hydride transfer and a proton transfer that seem to take place concertedly. The degree ofasynchronicity and the energy barrier depend on the particular starting reactant valley. In order to estimaterate constants we used a free energy perturbation technique adapted to follow the intrinsic reactioncoordinate for several possible reaction paths. Tunneling effects are also obtained with a slightly modifiedversion of the ensemble-averaged variational transition state theory with multidimensional tunnelingcontributions. According to our results the closure of the active site by means of a flexible loop can lead tothe formation of different reactant complexes displaying different features in the disposition of some keyresidues (such as Arg109), interactions with the substrate and number of water molecules in the activesite. The chemical step of the reaction takes place with a different reaction rate from each of thesecomplexes. Finally, primary kinetic isotope effects for replacement of the transferring hydrogen of the cofactorwith a deuteride are in good agreement with experimental observations, thus validating our methodology.