Study On Regulation Of Arginine In Saccharomyces Cerevisiae To Reduce Ethyl Carbamate In Wine

Ethyl carbamate(EC)as a byproduct of fermented foods and beverage is a potential carcinogen for humans.The EC is mainly derived from the reaction of urea and ethanol produced by yeast metabolism during fermentation in wine.To reduce the production of EC in wine,this study took the diploid industrial Saccharomyces cerevisiae YZ11(knockout one CAR1 allele)as the starting strain,and modified the strain from the urea precursor arginine transport and synthesis pathway.The precursor of EC,urea,is mostly produced by the decomposition of arginine during the fermentation process of S.cerevisiae.To reduce the intracellular arginine content,the genes encoding arginine permease(CAN1)and amino acid permease(GAP1),were respectively knocked out in this study.The results of wine fermentation showed that the EC production of the recombinant strains YC11 and YC21 decreased by 23.2% and 58.11%,respectively,compared with the starting strain YZ11;the EC production of recombinant strains YG11 and YG21 decreased by 66.88% and 61.04%,respectively,compared with the starting strain YZ11.No effects on the growth performance or other conditions of the recombinant strains occurred.The recombinant strains YA11 and YA21 were obtained by knockout of ARG1,the gene encoding arginine synthase,a key enzyme in arginine synthesis pathway.The results showed that the EC production of recombinant strains YA11 and YA21 decreased by14.29% and 42.28%,respectively,compared with the starting strain YZ11.No effects on the growth performance or other conditions of the recombinant strains occurred.In this study,recombinant strain YV11 were obtained by overexpressed VBA2 which the gene encoding basic amino acid transport protein in the tonoplast membrane.Using the recombinant strain for wine fermentation,the EC production of YV11 was reduced by52.94% compared with the starting strain YZ11.No effects on the growth performance or other conditions of the recombinant strains occurred.The combined transformation of CAN1 and GAP1,the results of wine fermentation showed that the EC production of recombinant strains YCG11 and YCG21 was reduced by52.43% and 56.35%,respectively,compared with the original strain YZ11.The recombinant yeast YVA11 obtained by knocking out the argininosuccinate synthase encoding gene ARG1 and overexpressing the basic amino acid transport protein encoding gene VBA2 located in the tonoplast had a 68% reduction in EC production in the wine fermentation experiment compared with the starting strain YZ11.No effects on the growth performance or other conditions of the recombinant strains occurred.