Metabolically engineered Saccharomyces cerevisiae for branched-chain ester productions

• Branched-chain esters are produced by the engineered yeast.
• Mitochondrial isobutanol biosynthetic pathway is used for ester synthesis.
• Alcohol acyltransferase is functional in yeast mitochondria.
• Segmentation the pathway into two compartments increased the ester yield.
• Subcellular metabolic engineering offers an effective way for ester synthesis.

Medium branched-chain esters can be used not only as a biofuel but are also useful chemicals with various industrial applications. The development of economically feasible and environment friendly bio-based fuels requires efficient cell factories capable of producing desired products in high yield. Herein, we sought to use a number of strategies to engineer Saccharomyces cerevisiae for high-level production of branched-chain esters. Mitochondrion-based expression of ATF1 gene in a base strain with an overexpressed valine biosynthetic pathway together with expression of mitochondrion-relocalized α-ketoacid decarboxylase (encoded by ARO10) and alcohol dehydrogenase (encoded by ADH7) not only produced isobutyl acetate, but also 3-methyl-1-butyl acetate and 2-methyl-1-butyl acetate. Further segmentation of the downstream esterification step into the cytosol to utilize the cytosolic acetyl-CoA pool for acetyltransferase (ATF)-mediated condensation enabled an additional fold improvement of ester productions. The best titre attained in the present study is 260.2 mg/L isobutyl acetate, 296.1 mg/L 3-methyl-1-butyl acetate and 289.6 mg/L 2-methyl-1-butyl acetate.