Cloning and characterization of phenolic acid decarboxylase responsible for aromatic volatile phenols production in Paocai based on metatranscriptomics

Sichuan Paocai is a kind of fermented vegetable with rich microorganism and enzyme resources. Based on our previous metatranscriptomics and flavor-oriented metabolomics studies, three potential phenolic acid decarboxylases LvPAD, DhFDC1 and LvUbiD were mined from environmental RNA of industrial Sichuan Paocai, and their catalytic function were verified by heterologous expression. LvPAD, which was derived from Lactobacillus vermolensis and had the highest catalytic activity among the three enzymes, was selected for further purification and characterization. It was optimally active at pH 6.0 and 35 degrees C, had good thermal stability and pH tolerance, and did not require specific cofactors. Kinetic parameters showed that LvPAD could catalyze the decarboxylation of p-coumaric acid, ferulic acid and caffeic acid with the kcat/Km values of 460.6 & PLUSMN; 0.1, 23.7 & PLUSMN; 0.09 and 118.2 & PLUSMN; 0.1 mM-1 s-1, respectively, and the products of the first two substrates were 4-vinylphenol and 4-vinylguaiacol with aromatic characteristics. Moreover, LvPAD was successfully used to preliminarily strengthen the vinylphenol aroma of industrial Paocai, which could shorten the fermentation time to a certain extent in practical production. Therefore, this work not only validated the important role of LvPAD in the production of aromatic phenols in Paocai, but also facilitated the mining of more enzymes from Paocai to improve the flavor and production process in the future.