The First Experimental Test of the Hypothesis that Enzymes Have Evolved To Enhance Hydrogen Tunneling

Doll, Kenneth M.; Bender, Bruce R.; Finke, Richard G.

doi:10.1021/ja030120h

Cited by 96 publications

(82 citation statements)

References 71 publications

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“…The aggregate data for SLO-1 that include huge primary deuterium kinetic isotope effects for WT-enzyme and all mutants studied thus far, an unexpectedly small temperature dependence of the size of the isotope effects with WT-enzyme, and highly temperature dependent KIEs for the distal mutations at position 553 provide some of the strongest evidence in support of a full tunneling process in an enzyme reaction. Whereas the importance of hydrogen tunneling has increasingly achieved wide acceptance, a discussion has ensued regarding the extent to which enzymes catalyze the tunneling event, i.e., whether it may be expected that the extent of tunneling will be the same or different in relation to a comparable reaction in solution (29)(30)(31). One approach to address this question is to study the properties of a model reaction in solution.…”

Section: Distinguishing the Types Of Heavy Atom Motions That Impact H-mentioning

confidence: 99%

Enzyme structure and dynamics affect hydrogen tunneling: The impact of a remote side chain (I553) in soybean lipoxygenase-1

Meyer

Tomchick²,

Klinman

2008

Proc. Natl. Acad. Sci. U.S.A.

153

293

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This study examines the impact of a series of mutations at position 553 on the kinetic and structural properties of soybean lipoxygenase-1 (SLO-1). The previously uncharacterized mutants reported herein are I553L, I553V, and I553G. High-resolution x-ray studies of these mutants, together with the earlier studied I553A, show almost no structural change in relation to the WT-enzyme. By contrast, a progression in kinetic behavior occurs in which the decrease in the size of the side chain at position 553 leads to an increased importance of donor–acceptor distance sampling in the course of the hydrogen transfer process. These dynamical changes in behavior are interpreted in the context of two general classes of protein motions, preorganization and reorganization, with the latter including the distance sampling modes [Klinman JP (2006) Philos Trans R Soc London Ser B 361:1323–1331; Nagel Z, Klinman JP (2006) Chem Rev 106:3095–3118]. The aggregate data for SLO-1 show how judicious placement of hydrophobic side chains can influence enzyme catalysis via enhanced donor–acceptor hydrogenic wave function overlap.

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Section: Distinguishing the Types Of Heavy Atom Motions That Impact H-mentioning

confidence: 99%

Enzyme structure and dynamics affect hydrogen tunneling: The impact of a remote side chain (I553) in soybean lipoxygenase-1

Meyer

Tomchick²,

Klinman

2008

Proc. Natl. Acad. Sci. U.S.A.

153

293

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“…Although it is now becoming widely accepted that hydron tunnelling does indeed occur in some of these enzymes (e.g. Ball 2004;Liang & Klinman 2004), it is not clear how widespread it is, or how important the phenomenon may be to catalysis (Bahnson et al 1997;Doll et al 2003;Warshel 2003). To contribute to this discussion, we have studied the enzyme methylamine dehydrogenase (MADH,EC 1.4.99.3) with the aim of understanding its structural features which promote or enhance the tunnelling mechanism.…”

Section: Introductionmentioning

confidence: 99%

An analysis of reaction pathways for proton tunnelling in methylamine dehydrogenase

Núñez

Tresadern

Hillier

et al. 2006

Phil. Trans. R. Soc. B

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Computational methods have now become a valuable tool to understand the way in which enzymes catalyse chemical reactions and to aid the interpretation of a diverse set of experimental data. This study focuses on the influence of the condensed-phase environment structure on proton transfer mechanisms, with an aim to understand how C-H bond cleavage is mediated in enzymatic reactions. We shall use a combination of molecular simulation, ab initio or semi-empirical quantum chemistry and semi-classical multidimensional tunnelling methods to consider the primary kinetic isotope effects of the enzyme methylamine dehydrogenase (MADH), with reference to an analogous application to triosephosphate isomerase. Analysis of potentially reactive conformations of the system, and correlation with experimental isotope effects, have highlighted that a quantum tunnelling mechanism in MADH may be modulated by specific amino acid residues, such as Asp428, Thr474 and Asp384.Keywords: proton tunnelling; methylamine dehydrogenase; enzyme catalysis; kinetic isotope effects; triosephosphate isomerase; quantum mechanical and molecular mechanical computational methods

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“…Although tantalizing, the relatively small difference in the observed KIEs is not sufficient to conclude that there is a greater tunneling contribution in the enzymatic process than that of the uncatalyzed reaction in water. A number of experimental and theoretical studies suggested that the extent of tunneling was the same in several enzymes as in model reactions in solution (2,9,10), whereas other experiments showing differential tunneling behaviors may be attributed to different reaction mechanisms (2,(11)(12)(13). Nuclear tunneling cannot be measured directly, and the best experimental diagnostic of tunneling is through measurement of kinetic isotope effects (14).…”

mentioning

confidence: 99%

Differential quantum tunneling contributions in nitroalkane oxidase catalyzed and the uncatalyzed proton transfer reaction

Major

Héroux

Orville

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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The proton transfer reaction between the substrate nitroethane and Asp-402 catalyzed by nitroalkane oxidase and the uncatalyzed process in water have been investigated using a path-integral free-energy perturbation method. Although the dominating effect in rate acceleration by the enzyme is the lowering of the quasiclassical free energy barrier, nuclear quantum effects also contribute to catalysis in nitroalkane oxidase. In particular, the overall nuclear quantum effects have greater contributions to lowering the classical barrier in the enzyme, and there is a larger difference in quantum effects between proton and deuteron transfer for the enzymatic reaction than that in water. Both experiment and computation show that primary KIEs are enhanced in the enzyme, and the computed Swain-Schaad exponent for the enzymatic reaction is exacerbated relative to that in the absence of the enzyme. In addition, the computed tunneling transmission coefficient is approximately three times greater for the enzyme reaction than the uncatalyzed reaction, and the origin of the difference may be attributed to a narrowing effect in the effective potentials for tunneling in the enzyme than that in aqueous solution.PI-FEP/UM simulations ͉ enzyme catalysis ͉ kinetic isotope effects ͉ X-ray structure A lthough the dominant factor in enzyme catalysis is the lowering of the quasiclassical free energy barrier of the enzymatic reaction in comparison with the uncatalyzed process (1-3), quantum mechanical tunneling has been recognized to also play a role in enzymatic hydrogen transfer reactions (2, 3). An intriguing, yet unanswered, question is whether enzymes have evolved to enhance tunneling to accelerate the reaction rate because quantum effects on rate acceleration are much smaller than hydrogen bonding and electrostatic stabilization of the transition state (1, 4, 5). Nevertheless, a small factor of two in rate enhancement can have important physiological impacts. Although it appears straightforward to address this question by comparing the enzymatic and the uncatalyzed reaction in solution, the difficulty is to design a model system that mimics exactly the same enzymatic reaction and mechanism. The present study examines the structure of nitroalkane oxidase (NAO) complex with nitroethane and kinetic isotope effects at the primary and secondary sites for the enzymatic and the uncatalyzed reaction. The computational findings are consistent with experimental data, suggesting that there is a differential tunneling effect for the proton transfer reaction in NAO and in water. Analysis of tunneling paths reveals that the enzyme reduces both the free energy of activation and the width of the effective potential, resulting in enhanced proton tunneling in the active site.The flavoenzyme NAO catalyzes the conversion of nitroalkanes to nitrite and aldehydes or ketones (Fig. S1) (6). The ␣-proton abstraction of the small substrate nitroethane by Asp-402 is rate-limiting, which is accelerated by a factor of 10 9 over the uncatalyzed reaction between n...

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The First Experimental Test of the Hypothesis that Enzymes Have Evolved To Enhance Hydrogen Tunneling

Cited by 96 publications

References 71 publications

Enzyme structure and dynamics affect hydrogen tunneling: The impact of a remote side chain (I553) in soybean lipoxygenase-1

Enzyme structure and dynamics affect hydrogen tunneling: The impact of a remote side chain (I553) in soybean lipoxygenase-1

An analysis of reaction pathways for proton tunnelling in methylamine dehydrogenase

Differential quantum tunneling contributions in nitroalkane oxidase catalyzed and the uncatalyzed proton transfer reaction

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