Recently, a team led by Dr. Yan Yan, a researcher at the South China Sea Institute of Oceanology, Chinese Academy of Sciences, in collaboration with Dr. Wang Wengui from the University of Jinan and Professor Tang Yi from the University of California, Los Angeles, completed a study on the biosynthesis of quadrane terrecyclic acid, a terpene compound derived from fungi. Their findings were published in "Chemical Science" and recognized as a featured paper of the week and a hot paper for 2024.
Terpenes are a class of compounds with 15 carbon atoms, characterized by various chain and ring structures composed of three isoprene units. They are widely distributed in plants, insects, microorganisms, and possess diverse biological activities, serving as important sources for pharmaceuticals, food, and cosmetics research. Terrecyclic acid, a quadrane terpene skeleton composed of fused cyclopentane and bicyclooctane rings, is a natural product with antibacterial and antitumor activities after oxidation modifications.
Due to the unique structure and biological activities of quadrane skeletons, research on their synthesis methods has attracted significant attention. However, the complex structure and cyclization patterns of these compounds result in multi-step syntheses with low yields, hindering the development of lead drugs. Mass production of quadrane natural products through synthetic biology methods is an effective strategy to address this challenge. Isotope tracing experiments suggest that the biosynthesis of quadrane skeletons may involve two distinct pathways, namely the Hirota pathway and the Coates pathway. While computational chemistry studies indicate a higher likelihood of the latter, the exact enzymatic mechanism behind the biosynthesis of quadrane skeletons remains to be confirmed.
By mining fungal genomes, the research team identified a biosynthetic gene cluster, ter, responsible for the synthesis of these compounds. This cluster includes a terpene cyclase (TerA), a P450 oxidation gene, a short-chain dehydrogenase gene (terC), and a major co-transporter superfamily gene. Through heterologous expression, in vivo feeding, and in vitro enzyme reactions, the team validated the functions of each gene in the cluster, confirming the biosynthetic pathway: TerA catalyzes the production of compound 25 from farnesyl pyrophosphate, which is then oxidized and carboxylated by the P450 enzyme TerB to yield compound 32, and finally, compound 32 is dehydrogenated by the enzyme TerC to form the final product 2.
Building upon the discovery of the novel TerA enzyme capable of converting farnesyl pyrophosphate to β-terrecyclene (25), the team confirmed through a series of site-directed mutagenesis studies and kinetic isotope effect analysis that the cyclization process follows the Coates pathway rather than the originally proposed Hirota pathway. This study unraveled the biosynthetic gene cluster for the antibacterial and antitumor quadrane terpene terrecyclic acid, elucidating the biosynthetic process of forming the quadrane skeleton through the action of the β-terrecyclene cyclase TerA, followed by oxidation and dehydrogenation modifications.
"The mechanism of quadrane skeleton formation was further confirmed through point mutations of the TerA protein and kinetic isotope effect analysis, solving decades-long mysteries surrounding the biosynthesis of this class of natural products," stated Dr. Yan Yan, the corresponding author of the paper. This research lays the foundation for further exploration of quadrane natural products from fungal databases using terpene cyclase TerA as a probe.
For more information on the research paper, visit: https://doi.org/10.1039/D4SC01208A
