Given the rapid development of genome mining in this decade, the substrate channel of paclitaxel might be identified in the near future. A robust microbial cell factory with gene dbat, encoding a key rate-limiting enzyme 10-deacetylbaccatin III-10-O-transferase (DBAT) in paclitaxel biosynthesis to synthesize the precursor baccatin III, will lay out a promising foundation for paclitaxel de novo synthesis. Here, we integrated gene dbat into the wild-type Escherichia coli BW25113 to construct strain BWD01. Yet, it was relatively unstable in baccatin III synthesis. Mutant gene dbat S189V with improved thermostability was screened out from a semi-rational mutation library of DBAT. When it was over-expressed in an engineered strain N05 with improved acetyl-CoA generation, combined with carbon source optimization of fermentation engineering, the production level of baccatin III was significantly increased. Using this combination, integrated strain N05S01 with mutant dbat S189V achieved a 10.50-fold increase in baccatin III production compared with original strain BWD01. Our findings suggest that the combination of protein engineering and metabolic engineering will become a promising strategy for paclitaxel production.
Recommended CitationHuang, Jia-Jun; Wei, Tao; Ye, Zhi-Wei; Zheng, Qian-Wang; Jiang, Bing-Hua; Han, Wen-Feng; Ye, An-Qi; Han, Pei-Yun; Guo, Li-Qiong; and Lin, Jun-Fang, "Microbial Cell Factory of Baccatin III Preparation in Escherichia coli by Increasing DBAT Thermostability and in vivo Acetyl-CoA Supply" (2022). Department of Pathology, Anatomy, and Cell Biology Faculty Papers. Paper 347.
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