Regiospecific modifications of naringenin for astragalin production in Escherichia coli

Sailesh Malla, Ramesh Prasad Pandey, Byung‐Gee Kim, Jae Kyung Sohng

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

We report the production of astragalin (AST) from regiospecific modifications of naringenin (NRN) in Escherichia coli BL21(DE3). The exogenously supplied NRN was converted into dihydrokaempferol (DHK) and then kaempferol (KMF) in the presence of flavanone‐3‐hydroxylase (f3h) and flavonone synthase (fls1) from Arabidopsis thaliana, respectively. KMF was further modified to produce AST by 3‐O‐glucosylation utilizing the endogeneous UDP‐glucose in presence of UGT78K1 from Glycine max. To increase the intracellular UDP‐glucose concentration by channeling the carbon flux toward UDP‐glucose at the branch point of glucose‐6‐phosphate (G6P), the chromosomal glucose phosphate isomerase (pgi) and D‐glucose‐6‐phosphate dehydrogenase (zwf) were knocked‐out in E. coli BL21(DE3). The two enzymes directly involved in the synthesis of UDP‐glucose from G6P, phosphoglucomutase (nfa44530) from Nocardia farcinia and glucose‐1‐phosphate uridylyltransferase (galU) from E. coli K12 were overexpressed, which successfully diverted the carbon flow from glycolysis to the synthesis of UDP‐glucose. Furthermore, to prevent the dissociation of UDP‐glucose into UDP and glucose, the UDP‐glucose hydrolase (ushA) was deleted. The E. coli ΔpgiΔzwfΔushA mutant harboring the UDP‐glucose biosynthetic pathway and the aforementioned genes for the regiospecific glucosylation produced 109.3 mg/L (244 µM) of AST representing 48.8% conversion from 500 µM of NRN in 60 h without any supplementation of extracellular UDP‐glucose. Biotechnol. Bioeng. 2013; 110:2525–2535. © 2013 Wiley Periodicals, Inc.
Original languageEnglish
JournalBiotechnology and Bioengineering
Volume110
Issue number9
Pages (from-to)2525-2535
ISSN0006-3592
DOIs
Publication statusPublished - 2013
Externally publishedYes

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