Canonical terpene synthases in arthropods: Intraphylum gene transfer.

Insects employ terpenoids for communication both within and between species. While terpene synthases derived from isoprenyl diphosphate synthase have been shown to catalyze terpenoid biosynthesis in some insects, canonical terpene synthases (TPS) commonly found in plants, fungi, and bacteria were previously unidentified in insects. This study reveals the presence of genes in insects, likely originating via horizontal gene transfer from noninsect arthropods.

Biosynthesis of a bacterial meroterpenoid reveals a non-canonical class II meroterpenoid cyclase.

Meroterpenoids are hybrid natural products that arise from the integration of terpenoid and non-terpenoid biosynthetic pathways. While the biosynthesis of fungal meroterpenoids typically follows a well-established sequence of prenylation, epoxidation, and cyclization, the pathways for bacterial perhydrophenanthrene meroterpenoids remain poorly understood. In this study, we report the construction of an engineered metabolic pathway in for the production of the bacterial meroterpenoid, atolypene A (1).

Early-stage biosynthesis of phenalinolactone diterpenoids involves sequential prenylation, epoxidation, and cyclization.

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 -perhydrophenanthrene core is constructed by sequential prenylation, epoxidation, and cyclization. The functions and timing of PlaT1-PlaT3 were assigned by comprehensive heterologous reconstitution.

Chemical Diversity of Aspergillus alliaceus Phenotypes: Discovery of Brominated Bianthrones with Activity against Triple-Negative Breast Cancer Cell Lines.

Marine-derived fungi have emerged as a source for novel metabolites with a broad range of bioactivities. However, accessing the full potential of fungi under standard laboratory conditions remains challenging. LC-MS-based metabolomics in combination with varied culture conditions is a fast and powerful tool to detect new metabolites.

Characterization of UbiA terpene synthases with a precursor overproduction system in Escherichia coli.

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.

Computation-guided scaffold exploration of 2,6-1,10-eunicellanes.

Eunicellane diterpenoids are a unique family of natural products containing a foundational 6/10-bicyclic framework and can be divided into two main classes, and , based on the configurations of their ring fusion at C1 and C10. Previous studies on two bacterial diterpene synthases, Bnd4 and AlbS, revealed that these enzymes form - and -eunicellane skeletons, respectively. Although the structures of these diterpenes only differed in their configuration at a single position, C1, they displayed distinct chemical and thermal reactivities.

Development of platensimycin, platencin, and platensilin overproducers by biosynthetic pathway engineering and fermentation medium optimization.

Unlabelled: The platensimycin (PTM), platencin (PTN), and platensilin (PTL) family of natural products continues to inspire the discovery of new chemistry, enzymology, and medicine. Engineered production of this emerging family of natural products, however, remains laborious due to the lack of practical systems to manipulate their biosynthesis in the native-producing Streptomyces platensis species. Here we report solving this technology gap by implementing a CRISPR-Cas9 system in S.

Class II terpene cyclases: structures, mechanisms, and engineering.

Covering: up to July 2023Terpene cyclases (TCs) catalyze some of the most complicated reactions in nature and are responsible for creating the skeletons of more than 95 000 terpenoid natural products. The canonical TCs are divided into two classes according to their structures, functions, and mechanisms. The class II TCs mediate acid-base-initiated cyclization reactions of isoprenoid diphosphates, terpenes without diphosphates (, squalene or oxidosqualene), and prenyl moieties on meroterpenes.

Cryptic Isomerization in Diterpene Biosynthesis and the Restoration of an Evolutionarily Defunct P450.

Biosynthetic modifications of the 6/10-bicyclic hydrocarbon skeletons of the eunicellane family of diterpenoids are unknown. We explored the biosynthesis of a bacterial -eunicellane natural product, albireticulone A (), and identified a novel isomerase that catalyzes cryptic isomerization in the biosynthetic pathway. We also assigned functions of two cytochromes P450 that oxidize the eunicellane skeleton, one of which was a naturally evolved non-functional P450 that, when genetically repaired, catalyzes allylic oxidation.

Cytochrome P450 Mediated Cyclization in Eunicellane Derived Diterpenoid Biosynthesis.

Terpene cyclization, one of the most complex chemical reactions in nature, is generally catalyzed by two classes of terpene cyclases (TCs). Cytochrome P450s that act as unexpected TC-like enzymes are known but are very rare. In this study, we genome-mined a cryptic bacterial terpenoid gene cluster, named ari, from the thermophilic actinomycete strain Amycolatopsis arida.

Biosynthesis, enzymology, and future of eunicellane diterpenoids.

Unlabelled: Eunicellane diterpenoids are a remarkable family of terpene natural products and have been of high interest for over five decades. Widely distributed in soft corals and rare in plants, eunicellanes were also recently identified in actinobacteria. These terpenoids have foundational 6/10-bicyclic frameworks that are frequently oxidized into structures containing transannular ether bridges.

First -eunicellane terpene synthase in bacteria.

Terpenoids are the largest family of natural products, but prokaryotes are vastly underrepresented in this chemical space. However, genomic supports vast untapped biosynthetic potential for terpenoids in bacteria. We discovered the first -eunicellane terpene synthase (TS), AlbS from NRRL B-1670, in nature.

Structure-guided product determination of the bacterial type II diterpene synthase Tpn2.

A grand challenge in terpene synthase (TS) enzymology is the ability to predict function from protein sequence. Given the limited number of characterized bacterial TSs and significant sequence diversities between them and their eukaryotic counterparts, this is currently impossible. To contribute towards understanding the sequence-structure-function relationships of type II bacterial TSs, we determined the structure of the terpentedienyl diphosphate synthase Tpn2 from Kitasatospora sp.

Discovery, Structure, and Mechanism of a Class II Sesquiterpene Cyclase.

Terpene cyclases (TCs), extraordinary enzymes that create the structural diversity seen in terpene natural products, are traditionally divided into two classes, class I and class II. Although the structural and mechanistic features of class I TCs are well-known, the corresponding details in class II counterparts have not been fully characterized. Here, we report the genome mining discovery and structural characterization of two class II sesquiterpene cyclases (STCs) from .

Mutation of the eunicellane synthase Bnd4 alters its product profile and expands its prenylation ability.

Bnd4 catalyzes the first committed step in the biosynthesis of the bacterial diterpenoid benditerpenoic acid and was the first eunicellane synthase identified from nature. We investigated the catalytic roles of the aromatic residues in the active site of Bnd4 through a series of mutation studies. These experiments revealed that large hydrophobic or aromatic side chains are required at F162 and Y197 for eunicellane formation and that selected mutations at W316 converted Bnd4 into a cembrane synthase.

Characterization and Structural Determination of CmnG-A, the Adenylation Domain That Activates the Nonproteinogenic Amino Acid Capreomycidine in Capreomycin Biosynthesis.

Capreomycidine (Cap) is a nonproteinogenic amino acid and building block of nonribosomal peptide (NRP) natural products. We report the formation and activation of Cap in capreomycin biosynthesis. CmnC and CmnD catalyzed hydroxylation and cyclization, respectively, of l-Arg to form l-Cap.

A roadmap for the functional annotation of protein families: a community perspective.

Over the last 25 years, biology has entered the genomic era and is becoming a science of 'big data'. Most interpretations of genomic analyses rely on accurate functional annotations of the proteins encoded by more than 500 000 genomes sequenced to date. By different estimates, only half the predicted sequenced proteins carry an accurate functional annotation, and this percentage varies drastically between different organismal lineages.

The Structure-Function Relationship of Human Bleomycin Hydrolase: Mutation of a Cysteine Protease into a Serine Protease.

Human bleomycin hydrolase (hBH) catalyzes deamidation of the anticancer drug bleomycins (BLM). This enzyme is involved in BLM detoxification and drug resistance. Herein, we report the putative BLM-binding site and catalytic mechanism of hBH.

Bacterial diterpene synthases prenylate small molecules.

The biosynthesis of terpenoid natural products begins with a carbocation-based cyclization or prenylation reaction. While these reactions are mechanistically similar, there are several families of enzymes, namely terpene synthases and prenyltransferases, that have evolved to specifically catalyze terpene cyclization or prenylation reactions. Here, we report that bacterial diterpene synthases, enzymes that are traditionally considered to be specific for cyclization, are capable of efficiently catalyzing both diterpene cyclization and the prenylation of small molecules.

The Natural Products Atlas 2.0: a database of microbially-derived natural products.

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.

Mechanistic Insights into the Formation of the 6,10-Bicyclic Eunicellane Skeleton by the Bacterial Diterpene Synthase Bnd4.

The eunicellane diterpenoids are a unique family of natural products seen in marine organisms, plants, and bacteria. We used a series of biochemical, bioinformatics, and theoretical experiments to investigate the mechanism of the first diterpene synthase known to form the eunicellane skeleton. Deuterium labeling studies and quantum chemical calculations support that Bnd4, from Streptomyces sp.

Discovery and Biosynthesis of a Structurally Dynamic Antibacterial Diterpenoid.

A new bicyclic diterpenoid, benditerpenoic acid, was isolated from soil-dwelling Streptomyces sp. (CL12-4). We sequenced the bacterial genome, identified the responsible biosynthetic gene cluster, verified the function of the terpene synthase, and heterologously produced the core diterpene.

Bacterial terpenome.

Covering: up to mid-2020 Terpenoids, also called isoprenoids, are the largest and most structurally diverse family of natural products. Found in all domains of life, there are over 80 000 known compounds. The majority of characterized terpenoids, which include some of the most well known, pharmaceutically relevant, and commercially valuable natural products, are produced by plants and fungi.