A Novel Semi-biosynthetic Route for Artemisinin Production Using Engineered Substrate-Promiscuous P450BM3
- 20 March 2009
- journal article
- research article
- Published by American Chemical Society (ACS) in ACS Chemical Biology
- Vol. 4 (4), 261-267
- https://doi.org/10.1021/cb900006h
Abstract
Production of fine chemicals from heterologous pathways in microbial hosts is frequently hindered by insufficient knowledge of the native metabolic pathway and its cognate enzymes; often the pathway is unresolved, and the enzymes lack detailed characterization. An alternative paradigm to using native pathways is de novo pathway design using well-characterized, substrate-promiscuous enzymes. We demonstrate this concept using P450BM3 from Bacillus megaterium. Using a computer model, we illustrate how key P450BM3 active site mutations enable binding of the non-native substrate amorphadiene. Incorporating these mutations into P450BM3 enabled the selective oxidation of amorphadiene artemisinic-11S,12-epoxide, at titers of 250 mg L−1 in E. coli. We also demonstrate high-yielding, selective transformations to dihydroartemisinic acid, the immediate precursor to the high-value antimalarial drug artemisinin.Keywords
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