Enthalpy and Entropy in Enzyme Catalysis - A Study of Lipase Enantioselectivity

Abstract: Biocatalysis has become a popular technique in organicsynthesis due to high activity and selectivity of enzymecatalyzed reactions. Enantioselectivity is a particularlyattractive enzyme property, which is utilized for theproduction of enantiopure substances. Determination of thetemperature dependence of enzyme enantioselectivity allows forthermodynamic analyses that reveal the contribution ofdifferential activation enthalpy, ?R-S?H?, and entropy, ?R-S?S?. In the present investigation the influenceof substrate structure, variations on enzyme structure and ofreaction media on the enantioselectivity ofCandida antarcticalipase B has been studied.The contribution of enthalpy, ?R-S?H?, and entropy, T?R-S?S?, to the differential free energy, ?R-S?G?, of kinetic resolutions ofsec-alcohols were of similar magnitude. Generally thetwo terms were counteracting, meaning that the enantiomerfavored by enthalpy was disfavored by entropy. 3-Hexanol was anexception where the preferred enantiomer was favored both byenthalpy and by entropy. Resolution of 1-bromo-2-butanolrevealed non-steric interactions to influence both ?R-S?H?and ?R-S?S?. Molecular modeling of the spatial freedom ofthe enzyme-substrate transition state indicated correlation tothe transition state entropy. The acyl chain length was shownto affect enantioselectivity in transesterifications of asec-alcohol.Point mutations in the active site were found to decrease orincrease enantioselectivity. The changes were caused by partlycompensatory changes in both ?R-S?H?and ?R-S?S?. Studies on single and double mutationvariants showed that the observed changes were notadditive.Enantioselectivity was strongly affected by the reactionmedia. Transesterifications of asec-alcohol catalyzed byCandida antarcticalipase B was studied in eight liquidorganic solvents and supercritical carbon dioxide. Acorrelation of enantioselectivity and the molecular volume ofthe solvent was found.Differential activation enthalpy, ?R-S?H?, and entropy, ?R-S?S?, display a compensatory nature. However thiscompensation is not perfect, which allows for modifications ofenantioselectivity. The components of the thermodynamicparameters are highly complex and interdependent but if theirroles are elucidated rational design of enantioselectiveenzymatic processes may be possible.Keywords:Biocatalysis, enzyme catalysis,Candida antarcticalipase B, enantioselectivity,enthalpy, entropy, CALB, enantiomeric ratio