Biosynthesis of Heptacyclic Duclauxins Requires Extensive Redox Modifications of the Phenalenone Aromatic Polyketide

Abstract

Duclauxins are dimeric and heptacyclic fungal polyketides with notable bioactivities. We characterized the cascade of redox transformations in the biosynthetic pathway of duclauxin from Talaromyces stipitatus. The redox reaction sequence is initiated by a cupin family dioxygenase DuxM that performs an oxidative cleavage of the peri-fused tricyclic phenalenone and affords a transient hemiketal-oxaphenalenone intermediate. Additional redox enzymes then morph the oxaphenoalenone into either an anhydride or a dihydrocoumarin-containing monomeric building block that is found in dimeric duxlauxins. Oxidative coupling between the monomers to form the initial C-C bond was shown to be catalyzed by a P450 monooxygenase, although the enzyme responsible for the second C-C bond formation was not found in the pathway. Collectively, the number and variety of redox enzymes used in the duclauxin pathway showcase Nature's strategy to generate structural complexity during natural product biosynthesis.

Aminoacyl sulfonamide assembly in SB-203208 biosynthesis

“…The multistep reaction from L-cysteine to 7 triggered by SbzM is intriguing, as such a cascade reaction initiated by a cupin oxidase has rarely been reported, except for the one catalyzed by the recently reported fungal cupin enzyme in phenalenone biosynthesis 41 . The cupin enzyme that oxidizes the sulfur of cysteine 42 and the one that abstracts the methylene proton at the α-position of dimethylsulfoniopropionate 29,30 were known; however, SbzM is a cupin-type cysteine decarboxylase.…”

Biosynthesis of a Tricyclo[6.2.2.02,7]dodecane System by a Berberine Bridge Enzyme‐Like Aldolase

“…Meanwhile, in post‐assembly tailoring steps, aldol reactions could increase the complexity of natural product scaffolds by introducing additional C−C linkages and stereocenters. Notable examples include the light‐sensitive phytotoxin hypocrellin A, the RNA synthesis inhibitor duclauxin and the phytotoxic stemphyloxin II ( 1 ) (Figure B). However, a dedicated enzyme that can catalyze such intramolecular stereoselective aldol reactions has not been reported in post‐modification steps, which is surprising given the prevalence of aldol reaction‐derived complex structures found in nature.…”

“…Herein,w ea ctivatedacryptic polyketide biosynthetic gene cluster that was upregulated in the fungal wheat pathogen Parastagonospora nodorum during plant infection;t his resulted in the productiono ft he phytotoxic stemphyloxin II (1). Through heterologous reconstruction of the biosynthetic pathway and in vitro assay by using cell-freel ysate from Aspergillus nidulans,w ed emonstrated that ab erberine bridge enzyme (BBE)-like protein SthB catalyzes an intramolecular aldol reaction to establish the bridgedt ricyclo[6.2.2.0 2,7 ]dodecane skeleton in the postassembly tailorings tep. The characterization of SthB as an aldolase enriches the catalytic toolboxo fc lassic reactions and the functional diversities of the BBE superfamily of enzymes.…”

“…For example, product template (PT) domains from fungal nonreducingp olyketides ynthases (NR-PKSs) are responsible for controlling the aldol cyclizations of poly-b-ketone intermediates during aromatic polyketide biosynthesis. [5] Meanwhile,i np ost-assembly tailorings teps, aldol reactionsc ould increase the complexity of natural product scaffolds by introducing additional CÀCl inkages and stereocenters.N otable examples include the light-sensitive phytotoxin hypocrellin A, [6] the RNA synthesis inhibitor duclauxin [7] and the phytotoxics temphyloxin II (1) [8] ( Figure 1B). However,adedicated enzymet hat can catalyze such intramolecular stereoselective aldol reactionsh as not been reported in post-modification steps, which is surprising given the prevalence of aldol reaction-derived complex structuresf ound in nature.…”

“…Finally,t he introduction of the second P450 gene sthD (not presenti nt he bet cluster) successfully transformed 5 into the end product stemphyloxinII( 1), thusa chieving the total biosynthesis of 1 in A. nidulans and confirming SthD catalyzed a hydroxylation at the C-15 methyl group on the side chain. Given that stemphyloxin I ( 7), ad ecalin with ah ydroxyl group at the same position, wasp reviously reporteda sanatural product from S. botryosum, [8] we examined whether the sth cluster can synthesize 7 in A. nidulans. Co-expression of sthA/E/ F/D (omitting sthB)r esulted in the production of an ovel metabolite 9.T he structure of 9 was characterized to contain an additional hydroxyl group at C-15 compared to 6,w hich was confirmedb yt he crucial 1 H-1 HC OSY correlationsofH 2 -15 (d H = 3.43 and 3.49 ppm)/H-12 (d H = 1.89 ppm) (Table S9, Supporting Information), and is herein named as dihydrostemphyloxin I.…”