Description of Hydrolase-Enantioselectivity Must be Based on the Actual Kinetic Mechanism: Analysis of the Kinetic Resolution of Glycidyl (2,3-Epoxy-1-Propyl) Butyrate by Pig Pancreas Lipase

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Abstract
The kinetic resolution of R,S-glycidyl (R,S-2,3-epoxy-1-propyl) butyrate catalyzed by pig pancreas lipase (PPL) was studied in monophasic and biphasic systems. The course of the resolution at ester concentrations exceeding 0.05 M or in the presence of R,S-glycidol (R,S-2,3-epoxy-1-propanol), could not be described by the equations derived for a one substrate enzyme with a minimal kinetic scheme (Chen et al., 1987). Trivial causes like heterogeneity in activity of the (crude) PPL preparation and equilibrium phenomena due to changing phase ratios could be excluded. An equation based on the kinetic mechanism of hydrolases, in which the acyl-enzyme intermediate is allowed to react with water as well as with the produced alcohol (quantified by the selectivity constant, α), was evaluated. All initial rate and conversion data could be adequately fitted with this equation, not only for PPL in the monophasic (free in solution) but also in the biphasic (adsorbed to the interface) systems where it exhibited better activity and enantioselectivity. Thus, the enantiomeric ratio (E) and α are intrinsic parameters of PPL, remaining constant during the course of the reaction. The correctness of the approach for the PPL-system indicates that description of enantioselectivity must be based on the actual kinetic mechanism of hydrolases.