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. 2010 Dec 8;110(12):PR41-67.
doi: 10.1021/cr1001035.

Update 1 of: Tunneling and dynamics in enzymatic hydride transfer

Zachary D Nagel  1 , Judith P Klinman
Affiliations

Update 1 of: Tunneling and dynamics in enzymatic hydride transfer

Zachary D Nagel et al. Chem Rev. .
No abstract available

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Figures

Figure 1
Figure 1
In the classical Marcus theory, the reactant and product potential surfaces are indicated by R and P, respectively, and their crossing point, ΔG‡, represents the transition state with a free energy of activation defined in terms of λ and ΔG° (eq 1 in text).
Figure 2
Figure 2
Comparison of the configuration of NAD+ based on X-ray structures of abortive complexes for mutants of HLADH with bound cofactor and trifluoroethanol. This structure reflects protein with two mutations, F93W and V203A, where F93W is the reference structure. Upon expansion of the active site behind the nicotinamide ring, this tilts away from the active site. Reproduced with permission from ref . Copyright 1997 National Academy of Sciences, U.S.A.
Figure 3
Figure 3
Relationship between the log of the bimolecular rate constant and the exponent (EXP in eq 6 in the text): (A) V203G; (B) V203A:F93W; (C) V203A; (D) V203L; (E) F93W; (F) L57F. Reproduced with permission from ref . Copyright 1997 National Academy of Sciences, U.S.A.
Figure 4
Figure 4
Time courses for hydrogen/deuterium exchange into ht-ADH, which has been digested into 21 peptides. The five peptides of ht-ADH that show a transition in the same temperature range (20–40 °C) as that for the transition in tunneling behavior (30 °C) are shown: 10 °C (closed circles); 20 °C (open circles); 40 °C (closed triangles); 55 °C (open triangles); 65 °C (closed squares). In part f is shown the temperature dependence of the weighted average rate constant for H/D exchange at each temperature for peptides 1–4 and 7. Reproduced with permission from ref . Copyright 2004 National Academy of Sciences, U.S.A.
Figure 5
Figure 5
(A) Structural representation of the peptides showing a temperature-dependent transition in H/D exchange behavior between 20 and 40 °C (cf. Figure 4): peptide 1, red; peptide 2, orange; peptide 3, purple; peptide 4, purple; peptide 7, red. (B) Those peptides showing a transition in H/D exchange between 40 and 55 °C (orange, red, and purple). Reproduced with permission from ref . Copyright 2004 National Academy of Sciences, U.S.A.
Figure 6
Figure 6
One-dimensional barrier penetration model invoked in the context of a tunneling correction. The lighter isotopes tunnel lower down in the barrier, generating elevated isotope effects, differences in enthalpies of activation that exceed the semiclassical limit, and values for AH/AD ≪ 1.
Figure 7
Figure 7
An expanded Marcus model applied to hydrogen transfer. The nuclear coordinates are represented by Q, the donor–acceptor (gating) coordinate is represented by rx, and ro is the initial equilibrium distance. Panel A is analogous to classical Marcus theory in that heavy nuclear reorganization (the free energy for this process is represented by the heavy lines as a function of progress along the coordinate Q) is required before significant tunneling can take place. The main difference is that hydrogen (and not electron) wave function overlap determines the tunneling probability. At the transition state, corresponding to the intersection of the heavy lines, the hydrogenic potential surfaces are isoenergetic with respect to hydrogen transfer, so that transitions (tunneling) can take place between them in accord with the Franck–Condon principle. Panel B illustrates how, once this configuration is reached, hydrogen wave function overlap can be enhanced by bringing the donor and acceptor closer together. The ability of thermal fluctuations to compress the oscillator by an amount Δr below its equilibrium configuration ro will depend on the force constant describing the curvature in the potential well. The distance rx is where the balance between interatomic repulsive forces and efficient wave function overlap is optimal. It is anticipated that, for many enzyme active sites, the well will be relatively stiff, so that the donor–acceptor distance remains near ro, and gating effects are minimized. Reproduced with permission from ref . Copyright 2005 Elsevier.
Figure 8
Figure 8
Two scenarios for enzyme active sites can be envisaged. According to (A) the protein dynamics and active site environment promote close approach between the H-donor and acceptor, leading to AH/AD > 1 and EXP > 3.26. For a “compromised” active site, a greater initial distance between the donor and acceptor atoms generates AH/AD < 1 and EXP ≈ 3.26.
Figure 9
Figure 9
Distinction between the types of motion expected to be critical to enzymatic H-transfer. On the left (A) is the “preorganization” term that involves a rapid sampling (ns–ms) of multiple conformations, preventing the active site from becoming trapped in a local minimum with a less optimal arrangement of bound substrates. On the right (B) is the subsequent “reorganization” of bound substrates. This is expected to involve even more rapid sampling (fs–ns) of states that differ with regard to the relative energy levels of reactant and product and the distances between reactants. The temperature dependence of the isotope effect comes primarily from the distance-sampling component of the reorganization term. When distance sampling has been minimized, as a result of effective preorganization events, the isotope effect can appear temperature-independent.
Figure 10
Figure 10
Native enzymes are able to transiently sample conformers in which a short equilibrium donor–acceptor distance r0 supports tunneling of both hydrogen and deuterium with little or no further distance sampling.
Figure 11
Figure 11
Kinetic mechanism of DHFR at 25 °C, where rate constants pertain to the protonated enzyme. The kinetic species represented by the abbreviations are as follows: N, NADP+; NH, NADPH; H2F, dihydrofolate (referred to as DHF in the text); H4F, tetrahydrofolate (referred to as THF in the text). Reproduced with permission from ref . Copyright 2006 Annual Reviews.
Figure 12
Figure 12
X-ray structure of E. coli dihydrofolate reductase modeled to be complexed with NADPH and DHF(H2F). (A) indicates mobile loops in gray, and (B) shows the location of conserved residues (red) and specifically G121 and M42 (red spheres). The black sphere indicates the position of hydride in the active site. Reproduced with permission from ref . Copyright 2002 American Chemical Society.
Figure 13
Figure 13
Hydrophobic residues at the active site of ec-DHFR that are proposed to control the relative position of NAD+ and substrate. Side chains of residues 14 and 94 are shown in red spheres, NAD in yellow, and folate in black. Reproduced with permission from ref . Copyright 2004 American Chemical Society.
Figure 14
Figure 14
Comparison of X-ray structures of homologous DHFRs. (A) Cα overlay for bs-DHFR (blue) and tm-DHFR (red) and (B) overlay for bs-DHFR (blue) and ec-DHFR (green). Reproduced with permission from ref . Copyright 2005 American Chemical Society.
Figure 15
Figure 15
Driving force dependence of the observed rate kcat/KM(O2) [left ordinate and red squares] and the kinetic isotope effect (kcat/KM(O2)) [right ordinate and blue circles].
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