Comparative analysis of the expression level of recombinant ginsenoside-transforming β-glucosidase in GRAS hosts and mass production of the ginsenoside Rh2-Mix

The ginsenoside Rh-2, a pharmaceutically active component of ginseng, is known to have anticancer and antitumor effects. However, white ginseng and red ginseng have extremely low concentrations of Rh-2 or Rh-2-Mix [20(S)-Rh-2, 20(R)-Rh-2, Rk(2), and Rh-3]. To enhance the production of food-grade ginsenoside Rh-2, an edible enzymatic bioconversion technique was developed adopting GRAS host strains. A beta-glucosidase (BglPm), which has ginsenoside conversion ability, was expressed in three GRAS host strains (Corynebacterium glutamicum, Saccharomyces cerevisiae and Lactococus lactis) by using a different vector system. Enzyme activity in these three GRAS hosts were 75.4%, 11.5%, and 9.3%, respectively, compared to that in the E. coli pGEX 4T-1 expression system. The highly expressed BglPm_C in C. glutamicum can effectively transform the ginsenoside Rg(3)-Mix [20(S)-Rg(3), 20(R)-Rg(3), Rk(1), Rg(5)] to Rh-2-Mix [20(S)-Rh-2, 20(R)-Rh-2, Rk(2), Rh-3] using a scaled-up biotransformation reaction, which was performed in a 10-L jar fermenter at pH 6.5/7.0 and 37 degrees C for 24 h. To our knowledge, this is the first report in which 50 g of PPD-Mix (Rb-1, Rb-2, Rb-3, R-c, and Rd) as a starting substrate was converted to ginsenoside Rg(3)-Mix by acid heat treatment and then 24.5-g Rh-2-Mix was obtained by enzymatic transformation of Rg(3)-Mix through by BglPm_C. Utilization of this enzymatic method adopting a GRAS host could be usefully exploited in the preparation of ginsenoside Rh-2-Mix in cosmetics, functional food, and pharmaceutical industries, thereby replacing the E. coli expression system.