Bioconversion of major ginsenosides Rg(1) to minor ginsenoside F-1 using novel recombinant ginsenoside hydrolyzing glycosidase cloned from Sanguibacter keddieii and enzyme characterization

Abstract

This study focuses on the cloning, expression, and characterization of recombinant ginsenoside hydrolyzing glycosidase from Sanguibacter keddieii in order to biotransform ginsenosides efficiently. The gene, termed bglSk, consists of 1857 bp and revealed significant homology to that of glycoside hydrolase family 3. The enzyme was over-expressed in Escherichia coli BL21 (DE3) using a GST-fused pGEX 4T-1 vector system. The over-expressed recombinant enzymes could convert six major ginsenosides Rb-1, Rb-2, Rc, Rd, Re and Rg(1) into more pharmacologically active rare ginsenosides such as C-Y, C-Mc, C-K, Rg(2)(S), and F-1. Especially, BglSk could completely convert the Rg1 into F1. The GST-fused BglSk was purified with GST bind agarose resin and then characterized. The kinetic parameters for beta-glucosidase had apparent K-m values of 0.456 +/- 0.009 and 0.167 +/- 0.003 mM and V-max values of 30.2 +/- 0.7 and 4.1 +/- 0.1 mu mol min(-1) mg of protein(-1) against p-nitrophenyl-beta-D-glucopyranoside and Rb-1, respectively. (C) 2012 Elsevier B.V. All rights reserved.