The Biosynthesis of Norsesquiterpene Aculenes Requires Three Cytochrome P450 Enzymes to Catalyze a Stepwise Demethylation Process

Abstract

Aculenes are a unique class of norsequiterpenes (C14) that are produced by Aspergillus aculeatus. The nordaucane skeleton in aculenes A–D may be derived from an ent‐daucane precursor through demethylation, however, the enzymes involved remain unexplored. We identified the biosynthetic gene cluster and characterized the biosynthetic pathway based on gene inactivation, feeding experiments, and heterologous reconstitution in Saccharomyces cerevisiae and Aspergillus oryzae. We discovered that three cytochrome P450 monoxygenases are required to catalyze the stepwise demethylation process. AneF converts the 12‐methyl group into a carboxylic acid and AneD installs the 10‐hydroxy group for later tautomerization and stabilization. Finally, AneG installs an electron‐withdrawing carbonyl group at the C‐2 position, which triggers C‐12 decarboxylation to yield the nordaucane skeleton. Additionally, a terpene cyclase (AneC) was found that forms a new product (dauca‐4,7‐diene).

Targeted Genome Mining Reveals the Biosynthetic Gene Clusters of Natural Product CYP51 Inhibitors

“…Hence, Rstn8 represents an example of the emerging class of single-module NRPS-like enzymes that performs esterification reactions. 27,34,35 Rstn8 displays strict substrate specificity toward glycine as no other natural amino acid was accepted (Figure S10). Rstn8 does not recognize 7 as a substrate, demonstrating that Rstn1-catalyzed methylation, possibly protecting the C4-OH, must precede the final esterification step.…”

Fungal Metabolite Asperaculane B Inhibits Malaria Infection and Transmission

“…Instead of repeating previously published work [ 12 , 14 , 15 ], we obtained a set of pure compounds ( Figure 6 b), including the pure asperaculane B ( Figure S1 ) from two laboratories, where the compounds were isolated from A. aculeatus in 2015 [ 12 ] and 2019 [ 14 ]. We determined the malaria transmission-blocking activity of these compounds with SMFA at 10 µg/mL and fed An.…”

“…( a ) The high-resolution mass spectrometry of the HPLC purified compound with a proton (236.1403 + 1.0078 = 237.1481). ( b ) Four purified and identified compounds from A. aculeatus from other research labs [ 12 , 14 ]. ( c ) The transmission-blocking activities of four compounds.…”

“…The active compound was isolated and determined to be asperaculane B. The structure of asperaculane B has resolved previously [ 12 , 14 ]. Notably, asperaculane B also inhibited asexual P. falciparum infection in blood while being nontoxic to human cells.…”

Genome Mining of Alkaloidal Terpenoids from a Hybrid Terpene and Nonribosomal Peptide Biosynthetic Pathway

“…8 Recent characterization of the aculene A biosynthetic pathway demonstrates such collaboration between the TC and a single module NRPS. 9 Based on this evidence, we believe there is significant potential in mining fungal TC/NRPS pathways for new NPs.…”