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Updated: Aug 6, 2025
A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
Tuning an efficient Escherichia coli whole-cell catalyst expressing l-pantolactone dehydrogenase for the biosynthesis
Fang-Ying Zhu1, Qing Yang2, Min Cao1
1National and Local Joint Engineering Research Center for Biomanufacturing of Chiral Chemicals, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Key Laboratory of Bioorganic Synthesis of Zhejiang Province, College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
Abstract:
d-(-)-Pantolactone (DPL) is a key intermediate for the production of d-(+)-pantothenate (vitamin B5). Deracemization of d,l-pantolactone (D,L-PL) through oxidizing l-(+)-pantolactone (LPL) to ketopantoyl lactone (KPL) and subsequently reducing KPL to DPL is a promising route for synthesizing DPL. Herein, a newly mined l-pantolactone dehydrogenase from Rhodococcus hoagie (RhoLPLDH) was used for the oxidative dehydrogenation of LPL. To alleviate inclusion bodies formed by membrane-bound RhoLPLDH intracellular expression in E. coli, strategies involving chaperone assistance and decreasing induction temperature were used to achieve RhoLPLDH soluble expression. To enhance its activity, directed evolution and hydrophilicity-based engineering yielded increased catalytic activity and thermostability. 1 M LPL was efficiently converted to KPL by engineering strain CM5 co-expressing RhoLPLDHL254I/V241I/I156L/F224Q/N164K and chaperone. A "two stages in one-pot" method was employed in deracemization of 1 M D,L-PL with 91.2% yield. These results demonstrated that CM5 catalyst exhibits great potential in enzyme cascade deracemization for the production of DPL.
