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The chemical logic associated with assembly of many bacterial terpenoids remains poorly understood. We focused our efforts on the early-stage biosynthesis of the phenalinolactone diterpenoids, demonstrating that the anti/anti/syn -perhydrophenanthrene core is constructed by sequential prenylation, epoxidation, and cyclization. The functions and timing of PlaT1-PlaT3 were assigned by comprehensive heterologous reconstitution. We illustrated that the UbiA prenyltransferase PlaT3 acts on geranylgeranyl diphosphate (GGPP) in the first step of phenalinolactone biosynthesis, prior to epoxidation by the flavin-dependent monooxygenase PlaT1 and cyclization by the type II terpene cyclase PlaT2. Finally, we isolated eight new-to-nature terpenoids, expanding the scope of the bacterial terpenome. The biosynthetic strategy employed in the assembly of the phenalinolactone core, with cyclization occurring after prenylation, is rare in bacteria and resembles fungal meroterpenoid biosynthesis. The findings presented here set the stage for future discovery, engineering, and enzymology efforts in bacterial meroterpenoids., Competing Interests: There are no conflicts of interest to declare., (This journal is © The Royal Society of Chemistry.)

Expression and purification of membrane-bound proteins remains a challenge and limits enzymology efforts, contributing to a substantial knowledge gap in the biochemical functions of many proteins found in nature. Accordingly, the study of bacterial UbiA terpene synthases (TSs) has been limited due to the experimental hurdles required to purify active enzymes for characterization in vitro. Previous work employed the use of microsomes or crude membrane fractions to test enzyme activity; however, these methods can be labor intensive, require access to an ultracentrifuge, or may not be suitable for all membrane-bound TSs. We detail here an alternative strategy for the in vivo expression and biochemical characterization of the membrane associated UbiA TSs by employing a precursor overproduction system in Escherichia coli., (Copyright © 2024. Published by Elsevier Inc.)

Within the natural products field there is an increasing emphasis on the study of compounds from microbial sources. This has been fuelled by interest in the central role that microorganisms play in mediating both interspecies interactions and host-microbe relationships. To support the study of natural products chemistry produced by microorganisms we released the Natural Products Atlas, a database of known microbial natural products structures, in 2019. This paper reports the release of a new version of the database which includes a full RESTful application programming interface (API), a new website framework, and an expanded database that includes 8128 new compounds, bringing the total to 32 552. In addition to these structural and content changes we have added full taxonomic descriptions for all microbial taxa and have added chemical ontology terms from both NP Classifier and ClassyFire. We have also performed manual curation to review all entries with incomplete configurational assignments and have integrated data from external resources, including CyanoMetDB. Finally, we have improved the user experience by updating the Overview dashboard and creating a dashboard for taxonomic origin. The database can be accessed via the new interactive website at https://www.npatlas.org., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)