Constructing Microbial Hosts for the Production of Benzoheterocyclic Derivatives

Abstract

Natural products containing benzoheterocyclic skeletons are widely found in plants and exhibit various pharmacological activities. To address the current limited availability of these compounds, we herein demonstrate the production of benzopyran, furanocoumarins, and pyranocoumarins in Streptomyces xiamenensis by employing prenyltransferases and two substrate-promiscuous enzymes, XimD and XimE. To avoid the degradation in S. xiamenensis, furanocoumarins and pyranocoumarins were also successfully produced in Escherichia coli. The production of linear furanocoumarins (marmesin) and angular pyranocoumarins (decursinol) reached 3.6 and 3.7 mg/L in shake flasks, respectively. To the best of our knowledge, this is the first report of the microbial production of the plant metabolites furanocoumarins and pyranocoumarins. Our study complements the missing link in the biosynthesis of pyranocoumarins by leveraging the catalytic promiscuity of microbial enzymes.

Computational Exploration of How Enzyme XimE Converts Natural S-Epoxide to Pyran and R-Epoxide to Furan

“…The regioselectivity of the XimE-catalyzed reaction is dependent on the chirality of the substrate, (2 R ,3 S )-pyran from the natural metabolite S- epoxide (2,5) and (2 R ,3 S )-furanocoumarin from the unnatural R -epoxy isomer . Indeed, pyran preference in xiamenmycin biosynthesis stems from the specific chirality of upstream epoxidation by the FAD-dependent oxidoreductase XimD …”

“…In the xiamenmycin biosynthesis of Streptomyces xiamenensis 318, the pyran ring of 3-chromanol xiamenmycin B is formed via S -epoxidation of an olefin precursor 3-geranyl-4-hydroxybenzoate by a FAD-dependent oxidoreductase (XimD). This S -epoxide is converted to a pyran by a SnoaL-like epoxide hydrolase (XimE) via epoxide-ring opening. − Because of the pharmaceutical potential of 3-chromanol skeletons, substrate ambiguity of XimE has been intensively explored with a wide scope of substrates, including halogen and hydroxyl substitution on the aromatic ring, benzoate ester, and other benzoheterocyclic compounds like 7-demethylsuberosin and osthenol . The cascade combination of XimD and XimE gives a ratio of pyranocoumarin to furanocoumarin of up to 50:1.…”

Synthetic and biosynthetic methods for selective cyclisations of 4,5-epoxy alcohols to tetrahydropyrans

“…112 In a follow-up study, this pair of substrate-promiscuous enzymes were further employed for combinatorial biosynthesis of a series of benzoheterocyclic derivatives with greater structural diversity. 113 3.2 Formation of THPs from 4,5-epoxy alcohols in which no EH has been identified 3.2.1 Ambruticin. The ambruticins are a group of myxobacterial polyketides produced by various Polyangium cellulosum and Sorangium cellulosum strains with potent antifungal activities against a broad range of pathogens.…”

“…136 In a recent study, Xu and co-workers engineered XimE from xiamenmycin biosynthesis to improve its catalytic activity for the preparation of angular pyranocoumarins by site-directed mutagenesis (Scheme 54). 113 Guided by the crystallographic structure of XimE and molecular docking, the Y119A mutant of XimE was generated. The proportion of THP product 226 to THF product 225 increased from 51.7% in wild-type XimE to 79.2% in the Y119A mutant.…”

“…The proportion of THP product 226 to THF product 225 increased from 51.7% in wild-type XimE to 79.2% in the Y119A mutant. 113 As further epoxide hydrolases are discovered, rational engineering of these enzymes to expand their substrate scope, enhance their catalytic activity and to possibly switch their regioselectivity in epoxide-opening process will be a fascinating area of research. For enzymes to be utilised in combination with organic synthesis, it will be necessary to optimise their stability, substrate scope and performance under a variety of conditions such as in the presence of organic solvents.…”