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Molecules 2017, 22(9), 1422;

Multienzyme Biosynthesis of Dihydroartemisinic Acid

Biotransformation Innovation Platform, Agency for Science Technology and Research, Singapore 138673, Singapore
Department of Biochemistry, National University of Singapore, Singapore 117598, Singapore
Authors to whom correspondence should be addressed.
Received: 13 August 2017 / Revised: 27 August 2017 / Accepted: 27 August 2017 / Published: 28 August 2017
(This article belongs to the Special Issue Multicomponent Reaction-Based Synthesis of Bioactive Molecules)
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One-pot multienzyme biosynthesis is an attractive method for producing complex, chiral bioactive compounds. It is advantageous over step-by-step synthesis, as it simplifies the process, reduces costs and often leads to higher yield due to the synergistic effects of enzymatic reactions. In this study, dihydroartemisinic acid (DHAA) pathway enzymes were overexpressed in Saccharomyces cerevisiae, and whole-cell biotransformation of amorpha-4,11-diene (AD) to DHAA was demonstrated. The first oxidation step by cytochrome P450 (CYP71AV1) is the main rate-limiting step, and a series of N-terminal truncation and transcriptional tuning improved the enzymatic activity. With the co-expression of artemisinic aldehyde dehydrogenase (ALDH1), which recycles NADPH, a significant 8-fold enhancement of DHAA production was observed. Subsequently, abiotic conditions were optimized to further enhance the productivity of the whole-cell biocatalysts. Collectively, approximately 230 mg/L DHAA was produced by the multi-step whole-cell reaction, a ~50% conversion from AD. This study illustrates the feasibility of producing bioactive compounds by in vitro one-pot multienzyme reactions. View Full-Text
Keywords: whole cell biocatalysis; CYP71AV1; dihydroartemisinic acid whole cell biocatalysis; CYP71AV1; dihydroartemisinic acid

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Chen, X.; Zhang, C.; Too, H.-P. Multienzyme Biosynthesis of Dihydroartemisinic Acid. Molecules 2017, 22, 1422.

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