Modulation of lipase properties in macro-aqueous systems by controlled enzyme immobilization: enantioselective hydrolysis of a chiral ester by immobilized Pseudomonas lipase

Lipase from Pseudomonas fluorescens (PFL) has been immobilized by using dif ferent immobilization protocols. The catalytic behavior of the different PF L derivatives in the hydrolytic resolution of f'ully soluble (R,S) 2-hydrox y l-phenyl butanoic acid ethyl ester (HPBE) in aqueous medium was analyzed. The soluble enzyme showed a significant but low enantioselectivity, hydrol yzing the S isomer more rapidly than the R-isomer (E = 7). The enzyme, immo bilized via a limited attachment to a long and flexible spacer arm, showed almost identical activity and specificity to the soluble enzyme. However, o ther derivatives, e.g. PFL adsorbed on supports covered by hydrophobic moie ties (octyl, decaoctyl), exhibited significant hyperactivation on immobiliz ation (approximately 7-fold). Simultaneously, the enantioselectivity of the PFL-immobilized enzyme was significantly improved (from E = 7 to E = 80). By using such derivatives, almost pure R eater isomer (e.e. > 99%) has been obtained after 55% hydrolysis of the racemic mixture of a solution of 10% (w/v) (R,S) HPBE. The derivatives could be used for 10 cycles without any s ignificant decrease in the activity of the biocatalyst. (C) 2001 Elsevier S cience Inc. All rights reserved.