Determination of half-reaction equilibrium in a ping-pong enzyme mechanism

Abstract

Substituted enzyme (or ping-pong) mechanisms usually involve enzymes that exist in two forms that alternate during the catalytic reaction. A method is described here for determining the position of the equilibrium of a half reaction in a ping-pong enzyme mechanism that is based on the kinetics of the burst reaction which occurs upon addition of reactants that recycle the enzyme from one form to another. The theoretical basis for the analysis is developed, and the method is applied to the half reaction of the aldimine form of aspartate transaminase with difluoro-oxaloacetate.

Stereoelectronic effects in the reaction of aromatic substrates catalysed by Halomonas elongata transaminase and its mutants

“…3,4 The mechanism underlying -transaminase activity has been generally assumed similar to that of aspartate transaminase; 5,6 only recently the reaction mechanism of Chromobacterium violaceum ω-transaminase has been investigated by means of density functional theory calculations and the energy profile for the conversion of (S)-1-phenylethylamine to acetophenone has been calculated. In this study the involved transition states and intermediates have been characterised.…”

“…The remarkable work of Codexis-Merck in evolving an amino transferase to achieve a "greener" synthesis of Sitagliptin, 1 highlighted the promising features of this class of enzymes as a biocatalytic tool for the manufacturing of chiral amines, 2 which are valuable building blocks for a wide range of bioactive compounds. 3,4 The mechanism underlying the ω-transaminase activity has been generally assumed similar to that of aspartate transaminase; 5,6 only recently the reaction mechanism of Chromobacterium violaceum ω-transaminase has been investigated by means of density functional theory calculations and the energy profile for the conversion of (S)-1-phenylethylamine to acetophenone has been calculated. In this study the involved transition states and intermediates have been characterised.…”