New insights into the function of plant tannase with promiscuous acyltransferase activity

SUMMARY Plant tannases (TAs) or tannin acyl hydrolases, a class of recently reported carboxylesterases in tannin‐rich plants, are involved in the degalloylation of two important groups of secondary metabolites: flavan‐3‐ol gallates and hydrolyzable tannins. In this paper, we have made new progress in studying the function of tea ( Camellia sinensis ) ( Cs ) TA—it is a hydrolase with promiscuous acyltransferase activity in vitro and in vivo and promotes the synthesis of simple galloyl glucoses and flavan‐3‐ol gallates in plants. We studied the functions of CsTA through enzyme analysis, protein mass spectrometry, and metabolic analysis of genetically modified plants. Firstly, CsTA was found to be not only a hydrolase but also an acyltransferase. In the two‐step catalytic reaction where CsTA hydrolyzes the galloylated compounds epigallocatechin‐3‐gallate or 1,2,3,4,6‐penta‐ O ‐galloyl‐β‐ d ‐glucose into their degalloylated forms, a long‐lived covalently bound Ser159‐linked galloyl–enzyme intermediate is also formed. Under nucleophilic attack, the galloyl group on the intermediate is transferred to the nucleophilic acyl acceptor (such as water, methanol, flavan‐3‐ols, and simple galloyl glucoses). Then, metabolic analysis suggested that transient overexpression of TAs in young strawberry ( Fragaria × ananassa ) fruits, young leaves of tea plants, and young leaves of Chinese bayberry ( Myrica rubra ) actually increased the total contents of simple galloyl glucoses and flavan‐3‐ol gallates. Overall, these findings provide new insights into the promiscuous acyltransferase activity of plant TA.