Enhancing ergothioneine production in Saccharomyces cerevisiae via protein fusion technology and gene integration optimization

Ergothioneine (EGT), a histidine derivative with diverse physiological functions, has broad applications in food, cosmetics, and pharmaceuticals. Fermentative production of EGT using recombinant Saccharomyces cerevisiae is widely considered the most cost-effective and direct method. However, no previous studies have explored improving EGT production in S. cerevisiae through protein fusion technology. In this study, we present the first successful construction of a fungal EGT biosynthesis pathway in S. cerevisiae , coupled with enzyme fusion techniques and optimization of gene copy integration. The engineered strain, SZA011, achieved a record EGT production of 122.5 mg/L under optimized shake flask fermentation conditions. Moreover, in a 10 L fed-batch fermentation system, SZA011 produced 626.24 mg/L of EGT. This study introduces a novel strategy for enhancing microbial EGT generation through protein and genetic engineering, potentially enabling industrial applications.