Reconstitution of Biosynthetic Machinery for the Synthesis of the Highly Elaborated Indole Diterpene Penitrem

Abstract

Penitrem A is one of the most elaborated members of the fungal indole diterpenes. Two separate penitrem gene clusters were identified using genomic and RNA sequencing data, and 13 out of 17 transformations in the penitrem biosynthesis were elucidated by heterologous reconstitution of the relevant genes. These reactions involve 1) a prenylation‐initiated cationic cyclization to install the bicyclo[3.2.0]heptane skeleton (PtmE), 2) a two‐step P450‐catalyzed oxidative processes forming the unique tricyclic penitrem skeleton (PtmK and PtmU), and 3) five sequential oxidative transformations (PtmKULNJ). Importantly, without conventional gene disruption, reconstitution of the biosynthetic machinery provided sufficient data to determine the pathway. It was thus demonstrated that the Aspergillus oryzae reconstitution system is a powerful method for studying the biosynthesis of complex natural products.

Orange, red, yellow: biosynthesis of azaphilone pigments in Monascus fungi

“…Similar enzymatic acetylation-elimination sequences that yield alkene functionalities have been described for the biosynthesis of tetronate and spirotetronate antibiotics in bacteria 42 , 43 and for the production of the indole diterpene penitrem in Penicillium spp. 44 …”

Hydroxylation with Unusual Stereoinversion Catalyzed by an FeII/2‐OG Dependent Oxidase and 3,6‐Diene‐2,5‐diketopiperazine Formation in the Biosynthesis of Brevianamide K

“…In typical hydroxylation reactions catalyzed by P450 or Fe II /2‐OG oxidases, the intermediate of porphyrin⋅ + ‐Fe IV =O in P450 or Fe IV =O in Fe II /2‐OG oxidase abstracts a hydrogen atom (H⋅) from the bound substrate (R 1 R 2 R 3 CH) to form porphyrin‐Fe IV ‐OH or Fe III ‐OH and the substrate radical (R 1 R 2 R 3 C⋅), then the hydroxyl radical (HO⋅) of porphyrin‐Fe IV ‐OH or Fe III ‐OH rebounds to the substrate radical, resulting in configuration retention in the product (Figures 4A and S20) [14] . To date, all reported enzymatic hydroxylation reactions of tertiary carbon centers in natural products have maintained the stereoconfiguration, such as some specific hydroxylation reactions during the biosynthesis of thaxtomin A, [17] bicyclomycin, [15e] novapikromycin, [18] penitrem A, [19] talaromyolide C [20] and ( S )‐3‐hydroxy‐L‐alloisoleucine [21] (Figure S21).…”

High-efficient production of mushroom polyketide compounds in a platform host Aspergillus oryzae

“…Transformation of A. oryzae NSAR1 [37] was carried out by the previously reported protoplast-polyethylene glycol method [38]. The plasmid pUARA2-herA was used for the first transformation to construct AO-herA.…”