Linking protein structure and dynamics to catalysis: the role of hydrogen tunnelling

Abstract

Early studies of enzyme-catalysed hydride transfer reactions indicated kinetic anomalies that were initially interpreted in the context of a 'tunnelling correction'. An alternate model for tunnelling emerged following studies of the hydrogen atom transfer catalysed by the enzyme soybean lipoxygenase. This invokes full tunnelling of all isotopes of hydrogen, with reaction barriers reflecting the heavy atom, environmental reorganization terms. Using the latter approach, we offer an integration of the aggregate data implicating hydrogen tunnelling in enzymes (i.e. deviations from Swain-Schaad relationships and the semi-classical temperature dependence of the hydrogen isotope effect). The impact of site-specific mutations of enzymes plays a critical role in our understanding of the factors that control tunnelling in enzyme reactions.

Figure 1

(a) The semi-classical view of H-transfer in which differences in rate among H, D and T arise from ground state vibrational levels. (b) A modification where tunnelling is allowed to occur below the top of the barrier. This is a tunnelling correction and is expected to be greater for H than D than T, given the trend in wavelengths as a function of the mass of the tunnelling particle.

Figure 2

A plot of the exponent relating the secondary isotope effect for hydride transfer in the reactions of the alcohol dehydrogenases as a function of the magnitude of the D/T isotope effect. These data are from table 2. The value for EXP of 13 is from the ht-ADH at 65 °C (table 4).

Figure 3

Predictions of the isotopic Arrhenius behaviour in the context of tunnelling correction, where greater movement through the barrier for H than D or T produces greater curvature in the Arrhenius plot.

Figure 4

A multidimensional picture of H-tunnelling in which the (a) reactant hydrogen coordinate samples different relative energy levels between reactant and product and different distances between the (b) donor and acceptor atoms (c) before proceeding to product.

Figure 5

Active site for soybean lipoxygenase illustrating the position of Leu754, Leu546 and Ile553 in relation to bound substrate. The reactive C–H of substrate is shown in black. The substrate has been modelled (Knapp et al. 2002) into the structure of the enzyme (Minor et al. 1996).