885610-08-2Relevant articles and documents
Highly enantioselective kinetic resolution of primary alcohols of the type Ph-X-CH(CH3)-CH2OH by Pseudomonas cepacia lipase: Effect of acyl chain length and solvent
Mezzetti, Alessandra,Keith, Curtis,Kazlauskas, Romas J.
, p. 3917 - 3924 (2007/10/03)
Although lipase from Pseudomonas cepacia (PCL) shows high enantioselectivity towards many secondary alcohols, it usually exhibits only low to moderate enantioselectivity towards primary alcohols. To increase this enantioselectivity, we optimised the reaction conditions for the PCL-catalysed hydrolysis of esters of three chiral primary alcohols: 2-methyl-3-phenyl-1- propanol 1, 2-phenoxy-1-propanol 2 and solketal 3. The enantioselectivity towards 1-acetate increased from E=16 to 38 upon changing the solvent from ethyl ether/phosphate buffer to 30% n-propanol in phosphate buffer and increased again to E ≥190 upon changing the substrate from 1-acetate to 1-heptanoate. The same changes increased the enantioselectivity towards alcohol 2 from E=17 to 70, but did not significantly increase the enantioselectivity towards alcohol 3. The best solvent was similar to the solvent used to crystallise the open form of PCL and likely stabilises the open form of PCL. This stabilisation may increase the enantioselectivity by removing kinetic contributions from a non-enantioselective lid-opening step. We determined the kinetic contribution of the lid-opening step by measuring the interfacial activation of PCL. The activation energy for the PCL-catalysed hydrolysis of ethyl acetate was at least 2.6 kcal/mol lower in the presence of a water-organic solvent interface.
Molecular basis for enantioselectivity of lipase from Pseudomonas cepacia toward primary alcohols. Modeling, kinetics, and chemical modification of Tyr29 to increase or decrease enantioselectivity
Tuomi, W. Victor,Kazlauskas, Romas J.
, p. 2638 - 2647 (2007/10/03)
Lipase from Pseudomonas cepacia (PCL) shows good enantioselectivity toward primary alcohols. An empirical rule can predict which enantiomer of a primary alcohol reacts faster, but there is no reliable strategy to increase the enantioselectivity. We used a combination of molecular modeling of lipase-transition state analogue complexes and kinetic measurements to identify the molecular basis of the enantioselectivity toward two primary alcohols: 2-methyl-3-phenyl-1-propanol, 1, and 2-phenoxy-1-propanol, 2. In hydrolysis of the acetate esters, PCL favors the (S)-enantiomer of both substrates (E = 16 and 17, respectively), but, due to changes in priorities of the substituents, the (S)-enantiomers of 1 and 2 have opposite shapes. Computer modeling of transition state analogues bound to PCL show that primary alcohols bind to PCL differently than secondary alcohols. Modeling and kinetics suggest that the enantioselectivity of PCL toward 1 comes from the binding of the methyl group at the stereocenter within a hydrophobic pocket for the fast-reacting enantiomer, but not for the slow-reacting enantiomer. On the other hand, the enantioselectivity toward 2 comes from an extra hydrogen bond between the phenoxy oxygen of the substrate to the phenolic OH of Tyr29. This hydrogen bond may slow release of the (R)-alcohol and thus account for the reversal of enantioselectvity. To decrease the enantioselectivity of PCL toward 2-acetate by a factor of 2 to E = 8, we eliminated the hydrogen bond by acetylation of the tyrosyl residues with N- acetylimidazole. To increase the enantioselectivity of PCL toward 2-acetate by a factor of 2 to E = 36, we increased the strength of the hydrogen bond by nitration of the tyrosyl residues with tetranitromethane. This is one of the first examples of a rationally designed modification of a lipase to increase enantioselectivity.
