Modulating acetate ester and higher alcohol production in Saccharomyces cerevisiae through the cofactor engineering

Flavor production by esters or by higher alcohols play a key role in the sensorial quality of fermented alcoholic beverages. In Saccharomyces cerevisiae cells, the syntheses of esters and higher alcohols are considerably influenced by intracellular CoA levels catalyzed by pantothenate kinase. In this work, we examined the effects of cofactor CoA and acetyl-CoA synthesis on the metabolism of esters and higher alcohols. Strains 12 alpha-BAP2 and 12 alpha+ATF1 where generated by deleting and overexpressing BAP2 (encoded branched-chain amino acid permease) and ATF1 (encoded alcohol acetyl transferases), respectively, in the parent 12 alpha strains. Then, 12 alpha-BAP2+CAB1 and 12 alpha-BAP2+CAB3 strains were obtained by overexpressing CAB1 (encoded pantothenate kinase Cab1) and CAB3 (encoded pantothenate kinase Cab3) in the 12 alpha-BAP2 strain, and 12 alpha-BAP2+CAB1+ATF1 and 12 alpha-BAP2+CAB3+ATF1 were generated by overexpressing ATF1 in the pantothenate kinase overexpression strains. The acetate ester level in 12 alpha-BAP2 was slightly changed relative to that in the control strain 12 alpha, whereas the acetate ester levels in 12 alpha-BAP2+CAB1, 12 alpha-BAP2+CAB3, 12 alpha-BAP2+CAB1+ATF1, and 12 alpha-BAP2+CAB3+ATF1 were distinctly increased (44-118% for ethyl acetate and 18-57% for isoamyl acetate). The levels of n-propanol, methyl-1-butanol, isopentanol, isobutanol, and phenethylol levels were changed and varied among the six engineered strains. The levels of acetate esters and higher alcohols can be modulated by changing the CoA and acetyl-CoA levels. The method proposed in this work supplies a practical means of breeding yeast strains by modulating acetate ester and higher alcohol production.