Biocatalytic synthesis of chiral intermediate of pregabalin with high substrate loading by a newly isolated Morgarella morganii ZJB-09203

Abstract

The chemoenzymatic process involving biocatalytic resolution of rac-2-carboxyethyl-3-cyano-5-methylhexanoic acid ethyl ester (CNDE, 1) has been the most competitive and attractive route for pregabalin. A new esterase-producing strain ZJB-09203, which exhibited high hydrolytic activity, excellent enantioselectivity, and diastereoselectivity towards CNDE, has been successfully isolated from soil samples with a pH indicator agar plate method. The isolate was identified as Morgarella morganii by the ATB system (ID 32 GN) and the 16S rDNA sequence. In order to suppress product inhibition during enzymatic hydrolysis of CNDE, an adsorptive biocatalytic process was developed by utilizing anion-exchange resin D201 as adsorbent for selective removal of (3S)-2-carboxyethyl-3-cyano-5-methylhexanoic acid (2) from the reaction medium. This approach allowed the substrate loading to be increased up to 1.5 M and the chiral intermediate 2 was produced in 682 mM, 45.3 % conversion, and 95 % ee. These results imply that M. morganii ZJB-09203 esterase is a promising biocatalyst in the development of chemenzymatic manufacturing process for pregabalin.