Correlating CH Bond Cleavage with Molybdenum Reduction in Xanthine Oxidase

Abstract

We have performed a computational study of substrate CÀH bond activation in enzymes of the XO family. The CÀH H-atom for all XO substrates studied is transferred to the terminal sulfido at the transition state with near neutral charge, and this is consistent with both Mo¼S p ! CÀH s* and CÀH s ! Mo¼S p* donorÀacceptor interactions activating the CÀH bond. A CÀH bond scission and Mo reduction appear to be highly correlated along the reaction coordinate for all XO substrates studied, with Mo reduction being a continuous and exponential function of CÀH bond breaking along the reaction coordinate.

Comparison of the Active-Site Design of Molybdenum Oxo-Transfer Enzymes by Quantum Mechanical Calculations

“…This high barrier is in accordance with the experimental observation that the desulfo form of XO is inactive, , although kinetic, spectroscopic, and crystallographic studies indicate that the related aldehyde oxidoreductase enzyme is active without the sulfido group . Kirk and co-workers have emphasized the role of the sulfido ligand for the proper electronic structure of the transition state for the hydride-transfer reaction. , Thus, we can conclude that the XO active site is carefully designed to make all three steps in the reaction mechanism possible: a proper acidity to make the initial proton transfer possible, a favorable net charge to enable the oxy transfer, and again a proper acidity to make the hydride transfer feasible. No native or designed enzyme model combines these properties as well as the XO model.…”

“…77 Kirk and coworkers have emphasized the role of the sulfido ligand for the proper electronic structure of the transition state for the hydride-transfer reaction. 78,79 Thus, we can conclude that the XO active site is carefully designed to make all three steps in the reaction mechanism possible: a proper acidity to make the initial proton transfer possible, a favorable net charge enable the oxy transfer, and again a proper acidity to make the hydride transfer feasible. No native or designed enzyme model combines these properties as well as the XO model.…”

The Mononuclear Molybdenum Enzymes

“…The key aspect that dictates the plane in which the chemistry occurs is the presence of a thiolate or sulfido ligand adjacent to the catalytically labile oxygen, and the availability of an equatorial in-plane S π bond that can accept the departing hydride from substrate. 275 …”

Xanthine oxidase–product complexes probe the importance of substrate/product orientation along the reaction coordinate

“…We have shown that the degree of Mo reduction along the reaction coordinate is strongly correlated to the degree of C substrate -H bond cleavage for a variety of XO/XDH substrates, including lumazine. [48] A longer C-H distance observed at the TS was shown to correlate with a greater percentage of Mo reduction at the TS. [48] We define the percent Mo reduction at the TS as the %Mo(xy) character in the TS redox orbital divided by the %Mo(xy) character in the reduced IM2 E-P complex.…”

“…[48] A longer C-H distance observed at the TS was shown to correlate with a greater percentage of Mo reduction at the TS. [48] We define the percent Mo reduction at the TS as the %Mo(xy) character in the TS redox orbital divided by the %Mo(xy) character in the reduced IM2 E-P complex. We find that the Mo(xy) character present in the TS redox orbital is ~28%, while that in the Mo(IV)-4-TV product HOMO is ~73%.…”