Engineering Saccharomyces cerevisiae for improving itaconic acid production

Abstract

Itaconic acid (ITA) is an unsaturated organic acid used in industrial production due to its versatility as a polymer building block. Engineering microbial cell factories for ITA biosynthesis from cost-effective and renewable raw materials has gained significant attention. Here, we performed combinatorial engineering using Saccharomyces cerevisiae to improve ITA production. First, exogenous cis-aconitic acid decarboxylase (CAD) was integrated into S. cerevisiae to construct the ITA-producing chassis. Then, the rate-limiting step was eliminated by changing the promoter that drives CAD expression to optimize ITA synthesis. A mitochondrial cis-aconitate transporter MTTA was also expressed to facilitate the transport of precursor, which resulted in an ITA titer of 244 mg/L. Furthermore, with overexpression of truncated citrate synthase tCIT2, an increased titer of 409 mg/L was obtained. Finally, the transport protein Qdr3 was overexpressed to enhance the export of ITA, resulting in a production of 578 mg/L in shake flask. In a 5-L bioreactor, the ITA titer reached 1.2 g/L, representing the highest reported level in S. cerevisiae. Overall, an advanced recombinant yeast strain was constructed for the efficient production of ITA via combinatorial metabolic engineering.

Keywords

Itaconic acid / Microbial cell factories / Saccharomyces cerevisiae / Metabolic engineering / Transport protein

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Funding

National Key Research and Development Program of China(2023YFF1104104)

National Natural Science Foundation of China(32100439)

China National Postdoctoral Program for Innovative Talents(BX20240145)

Jiangsu Funding Program for Excellent Postdoctoral Talent(2024ZB009)

RIGHTS & PERMISSIONS

Jiangnan University