Characterization of Regioselective Glycosyltransferase of Rhizobium Pusense JCM 16209T Useful for Resveratrol 4'-O-α-d-glucoside Production.

Enzymatic glycosylation is an industrially useful technique for improving the properties of compounds with hydroxy groups, and the biological activities of the resulting glycosides differ depending on the glycosylation position. Therefore, regioselective glycosyltransferases are required for precise synthesis of glycosides. We found that Rhizobium pusense JCM 16209T could catalyze the regioselective glycosylation of resveratrol. To identify the regioselective glycosyltransferase, two a-glucosidases of R. pusense JCM 16209T(RpG I and RpG II) were cloned and expressed in Escherichia coli. The molecular mass of purified recombinant RpG I and II was estimated to be 60 kDa by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE). RpG I showed strong glycosylation activity toward resveratrol with 4'selectivity of 98.3%. The enzyme activity was maximized at pH 8.0 and 50 degrees C, and enhanced in the presence of Cs+ and Li+ ions. The maximum molar yield of resveratrol 4'-O-a-glucoside from resveratrol reached 41.6% at 30 min, and the concentration of the product was 2.08 mmol L-1. Glycosylation activity was observed toward resveratrol as well as toward caffeic acid, ferulic acid, 6-gingerol, flavonoid, and isoflavonoid compounds with high regioselectivity, indicating that RpG I could glycosylate a wide range of substrates. To the best of our knowledge, there are few reports on microbial glycosyltransferases that are useful for regioselective glycosylation. This research could be the first step toward developing technologies for the precise synthesis of glycosides. (c) 2022, The Society for Biotechnology, Japan. All rights reserved.