Microbial synthesis of terephthalic acid via Saccharomyces cerevisiae cell factories

Terephthalic acid (TPA) is a key industrial chemical widely used in plastic films, bottle containers, pharmaceutical intermediates, and various other applications. Microbial synthesis of TPA has significant potential for sustainable development. In this study, a Saccharomyces cerevisiae cell factory was successfully designed to synthesize TPA using multiple engineering techniques. A genetically engineered S. cerevisiae strain was first constructed and used as the chassis by knocking out six alcohol dehydrogenases to block the branched metabolic pathways of intermediate metabolites such as p-tolualdehyde and 4-carboxybenzaldehyde. Screening of the integration loci helped to improve the titer of p-toluic acid in S. cerevisiae. Through several rounds of integration and enzyme fusion engineering, the heterogeneous TsaMB enzyme was successfully expressed in S. cerevisiae, enabling TPA biosynthesis via a well-planned two-stage biphasic fermentation strategy. Stepwise fermentation optimization revealed critical factors for increasing the TPA yield, with a 2.6-fold increase in the 4-carboxybenzyl alcohol titer and 33.5 % increase in the TPA titer by multiplying the copy number of tsaC gene. Ultimately, a S. cerevisiae cell factory achieved a TPA titer of 131.5 mg/L, corresponding to a conversion yield of 24.1 mol%. These results demonstrate the viability of microbial cell factory for TPA biosynthesis and provide new opportunities for the sustainable production of TPA.