Affiliation:
1. Department of General Surgery Jinshan District Central Hospital Affiliated Shanghai University of Medicine and Health Sciences Shanghai China
2. State Key Laboratory of Bioreactor Engineering, Newworld Institute of Biotechnology East China University of Science and Technology Shanghai China
3. School of Health Science and Engineering University of Shanghai for Science and Technology Shanghai China
4. Shanghai Key Laboratory of Molecular Imaging, School of Pharmacy Shanghai University of Medicine and Health Sciences Shanghai China
Abstract
AbstractErgothioneine (EGT) is a rare thiohistidine derivative with exceptional antioxidant properties. The blood level of EGT is considered highly reliable predictors for cardiovascular diseases and mortality, yet animals lack the ability to synthesize this compound. Free plasmids have been previously used to overexpress genes involved in the EGT biosynthetic pathway of Mycolicibacterium neoaurum. Here, we tentatively introduced a putative transporter gene mfsT1 into high‐copy plasmids and sharply increased the ratio of extracellular EGT concentration from 18.7% to 44.9%. Subsequently, an additional copy of egtABCDE, hisG, and mfsT1 was inserted into the genome with a site‐specific genomic integration tool of M. neoaurum, leading a 2.7 times increase in EGT production. Co‐enhancing the S‐adenosyl‐L‐methionine regeneration pathway, or alternatively, the integration of three copies of egtABCDE, hisG and mfsT1 into the genome further increased the total EGT yield by 16.1% (64.6 mg/L) and 21.7% (67.7 mg/L), respectively. After 168‐h cultivation, the highest titer reached 85.9 mg/L in the latter strain with three inserted copies. This study provided a solid foundation for genome engineering to increase the production of EGT in M. neoaurum.
Funder
National Natural Science Foundation of China