Extending the Structural Diversity of Labdane Diterpenoids from Marine-Derived Fungus Talaromyces sp. HDN151403 Using Heterologous Expression.

Heterologous biosynthesis has become an effective means to activate fungal silent biosynthetic gene clusters (BGCs) and efficiently utilize fungal genetic resources. Herein, thirteen labdane diterpene derivatives, including five undescribed ones named talarobicins A-E (3-7), were discovered via heterologous expression of a silent BGC (labd) in Aspergillus nidulans. Their structures with absolute configurations were elucidated using extensive MS and NMR spectroscopic methods, as well as electronic circular dichroism (ECD) calculations. These labdanes belong to four skeleton types, and talarobicin B (4) is the first 3,18-dinor-2,3:4,18-diseco-labdane diterpene with the cleavage of the C2-C3 bond in ring A and the decarboxylation at C-3 and C-18. Talarobicin B (4) represents the key intermediate in the biosynthesis of penioxalicin and compound 13. The combinatorial heterologous expression and feeding experiments revealed that the cytochrome P450 enzymes LabdC, LabdE, and LabdF were responsible for catalyzing various chemical reactions, such as oxidation, decarboxylation, and methylation. All of the compounds are noncytotoxic, and compounds 2 and 8 displayed inhibitory effects against methicillin-resistant coagulase-negative staphylococci (MRCNS) and Bacillus cereus.

Overexpression of Global Regulator SCrp Leads to the Discovery of New Angucyclines in Streptomyces sp. XS-16.

Six angucyclines including three unreported compounds (1-3) were isolated from Streptomyces sp. XS-16 by overexpressing the native global regulator of SCrp (cyclic AMP receptor). The structures were characterized based on nuclear magnetic resonance (NMR) and spectrometry analysis and assisted by electronic circular dichroism (ECD) calculations. All compounds were tested for their antitumor and antimicrobial activities, and compound 1 showed different inhibitory activities against various tumor cell lines with IC50 values ranging from 0.32 to 5.33 µM.

Meroterpenoids with Immunosuppressive Activity from Edible Fungus Craterellus odoratus.

Two unusual polyketide-sesquiterpene metabolites, craterodoratins T (1) and U (2), along with the known compound craterellin A (3), were isolated from the higher fungus Craterellus odoratus. The structures of isolated compounds were characterized based on nuclear magnetic resonance (NMR) and mass spectrum (MS) spectroscopic analysis, while the absolute configuration of the compounds was determined by theoretical NMR and electronic circular dichroism (ECD) calculations. Compound 1 possessed a rare structure with two aromatic groups. Compounds 1 and 3 showed immunosuppressive activity with IC50 values ranging from 5.516 to 19.953 µM.

Trichothecrotocins D-L, Antifungal Agents from a Potato-Associated Trichothecium crotocinigenum.

Seven new merosesquiterpenoids, trichothecrotocins D-J (1-7), two new trichothecene sesquiterpenoids, trichothecrotocins K (12) and L (13), and six known compounds (8-11, 14, and 15), were isolated from a potato-associated fungus, Trichothecium crotocinigenum. Compounds 5 and 6 were racemates which were further separated as pure enantiomers. Structures together with absolute configurations were established by extensive spectroscopic analysis, as well as quantum chemistry calculations on ECD and optical rotations. Compounds 1-4 are rare meroterpenoids featuring a seco-phenyl group, while 1 and 2 possessed a novel 6-6/5 fused ring system. Compounds 1-4, 8, 11, and 12 showed antifungal activity against four plant pathogens with MIC values of 8-128 µg/mL. It is suggested that the meroterpenoids produced by T. crotocinigenum may play an important role in the antifungal property of the fungus, thereby protecting the host plant, i.e., potato.

Antiviral and cytotoxic indole diterpenoids from the antarctic sponge-derived fungus Aspergillus candidus HDN15-152.

One new indole diterpenoid, ascandinine T (1), and three known analogues (2-4) were isolated from an Antarctic sponge-derived fungus Aspergillus candidus HDN15-152. The structures, including absolute configurations, were established based on NMR, HRESIMS, and electronic circular dichroism (ECD) calculations. All isolated compounds were tested for antiviral and anticancer activity. Compound 4 displayed antiviral activity against influenza A virus (IAV) of A/PR/8/34(H1N1) strain with an IC50 value of 39.2 µM, while compound 2 showed cytotoxicity against NCI-H446, NCI-H446/EP and L-02 cells with IC50 values ranging from 9.77 to 13.91 µM.

Exploring the Diverse Landscape of Fungal Cytochrome P450-Catalyzed Regio- and Stereoselective Dimerization of Diketopiperazines.

Dimeric indole-containing diketopiperazines (di-DKPs) are a diverse group of natural products produced through cytochrome P450-catalyzed C-C or C-N coupling reactions. The regio- and stereoselectivity of these reactions plays a significant role in the structural diversity of di-DKPs. Despite their pivotal role, the mechanisms governing the selectivity in fungi are not fully understood. Employing bioinformatics analysis and heterologous expression experiments, five undescribed P450 enzymes (AmiP450, AcrP450, AtP450, AcP450, and AtuP450) responsible for the regio- and stereoselective dimerization of diketopiperazines (DKPs) in fungi are identified. The function of these P450s is consistent with phylogenetic analysis, highlighting their dominant role in controlling the dimerization modes. Combinatorial biosynthesis-based pathway reconstitution of non-native gene clusters expands the chemical space of fungal di-DKPs and reveals that the regioselectivity is influenced by the substrate. Furthermore, multiple sequence alignment and molecular docking of these enzymes demonstrate a C-terminal variable region near the substrate tunnel entrance in AtuP450 that is crucial for its regioselectivity. These findings not only reveal the secret of fungal di-DKPs diversity but also deepen understanding of the mechanisms and catalytic specificity involved in P450-catalyzed dimerization reactions.

Biosynthesis of Atypical Angucyclines Unveils New Ring Rearrangement Reactions Catalyzed by Flavoprotein Monooxygenases.

Six new angucycline structures, including spirocyclione A (1), which contains an unusual oxaspiro[5.5]undecane architecture, and its ring-A-cleaved product spirocyclione B (2), were discovered by heterologous expression of a type II polyketide biosynthetic gene cluster captured from a marine actinomycete strain Streptomyces sp. HDN155000. Three flavoprotein monooxygenases are confirmed to be responsible for the oxidative carbon skeleton rearrangements in the biosynthesis of compounds 1 and 2. The obtained compounds showed promising cytotoxicity against different types of cancer cells.

Diterpenes Specially Produced by Fungi: Structures, Biological Activities, and Biosynthesis (2010-2020).

Fungi have traditionally been a very rewarding source of biologically active natural products, while diterpenoids from fungi, such as the cyathane-type diterpenoids from Cyathus and Hericium sp., the fusicoccane-type diterpenoids from Fusicoccum and Alternaria sp., the guanacastane-type diterpenoids from Coprinus and Cercospora sp., and the harziene-type diterpenoids from Trichoderma sp., often represent unique carbon skeletons as well as diverse biological functions. The abundances of novel skeletons, biological activities, and biosynthetic pathways present new opportunities for drug discovery, genome mining, and enzymology. In addition, diterpenoids peculiar to fungi also reveal the possibility of differing biological evolution, although they have similar biosynthetic pathways. In this review, we provide an overview about the structures, biological activities, evolution, organic synthesis, and biosynthesis of diterpenoids that have been specially produced by fungi from 2010 to 2020. We hope this review provides timely illumination and beneficial guidance for future research works of scholars who are interested in this area.

Sesquiterpenoids Specially Produced by Fungi: Structures, Biological Activities, Chemical and Biosynthesis (2015-2020).

Fungi are widely distributed in the terrestrial environment, freshwater, and marine habitat. Only approximately 100,000 of these have been classified although there are about 5.1 million characteristic fungi all over the world. These eukaryotic microbes produce specialized metabolites and participate in a variety of ecological functions, such as quorum detection, chemical defense, allelopathy, and maintenance of symbiosis. Fungi therefore remain an important resource for the screening and discovery of biologically active natural products. Sesquiterpenoids are arguably the richest natural products from plants and micro-organisms. The rearrangement of the 15 high-ductility carbons gave rise to a large number of different skeletons. At the same time, abundant structural variations lead to a diversification of biological activity. This review examines the isolation, structural determination, bioactivities, and synthesis of sesquiterpenoids that were specially produced by fungi over the past five years (2015-2020).

Immunosuppressive Sesquiterpenoids from the Edible Mushroom Craterellus odoratus.

The aim of this work was to comprehensively understand the chemical constituents of the edible mushroom Craterellus ordoratus and their bioactivity. A chemical investigation on this mushroom led to the isolation of 23 sesquiterpenoids including eighteen previously undescribed bergamotane sesquiterpenes, craterodoratins A-R (1-18), and one new victoxinine derivative, craterodoratin S (19). The new structures were elucidated by detailed interpretation of spectrometric data, theoretical nuclear magnetic resonance (NMR) and electronic circular dichroism (ECD) calculations, and single-crystal X-ray crystallographic analysis. Compounds 1 and 2 possess a ring-rearranged carbon skeleton. Compounds 3, 10, 12-15, 19, 20 and 23 exhibit potent inhibitory activity against the lipopolysaccharide (LPS)-induced proliferation of B lymphocyte cells with the IC50 values ranging from 0.67 to 22.68 µM. Compounds 17 and 20 inhibit the concanavalin A (ConA)-induced proliferation of T lymphocyte cell with IC50 values of 31.50 and 0.98 µM, respectively. It is suggested that C. ordoratus is a good source for bergamotane sesquiterpenoids, and their immunosuppressive activity was reported for the first time. This research is conducive to the further development and utilization of C. ordoratus.

Chemical constituents and their biological activities from the mushroom Pyropolyporus fomentarius.

The chemical constituents and their biological activities of the mushroom Pyropolyporus fomentarius were investigated in this study. Two previously undescribed pentacyclic lupane-type triterpenes, 3-formyloxybetulin and 3-formyloxybetulinic acid, two rare degraded ergosterols, pyropolincisterols A and B, along with ten known triterpenoids and four known ergosterols were isolated from the fruiting bodies of P. fomentarius. Their chemical structures were determined using a combination of spectroscopic analysis. Nine compounds exhibited certain cytotoxicities to human cancer cell lines, while polyporenic acid showed significant cytotoxicities to SMMC-7721 and A-549 with IC50 values less than 10 µM. Four compounds showed inhibitory activities against nitric oxide (NO) production in LPS-activated RAW264.7 macrophages with IC50 values of 36.3, 25.1, 21.4, and 34.2 µM, respectively. The results of this assessment suggested that the lanostane triterpenoids and ergosterols in fruiting bodies of P. fomentarius played key roles in its folk usages.

Melodinines Y1-Y4, four monoterpene indole alkaloids from Melodinus henryi.

Melodinines Y1-Y4 (1-4), four undescribed alkaloids were isolated from Melodinus henryi. Their structures were elucidated by extensive NMR, mass spectroscopic analyses, theoretical NMR and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 are the first examples of bisindole alkaloids possessing an eburnamine-leuconoxine combination. Compound 3 is a rare 2,3-seco pleiocarpamine type monoterpene indole alkaloid. Compound 1 showed cytotoxic activities against six human cancer cell lines with IC50 values of 0.5-15.2 µM.

Illudane Sesquiterpenoids from Edible Mushroom Agrocybe salicacola and Their Bioactivities.

To comprehensively understand the chemical constituents of the edible mushroom Agrocybe salicacola and their biological functions, a phytochemical separation of the cultural broth of A. salicacola led to the isolation of four new illudane sesquiterpenoids, agrocybins H-K (1-4), along with 10 known analogues (5-14). Compounds 2-4 were racemates of which 2 and 3 were further separated into single enantiomers as 2a/2b and 3a/3b. All new structures with absolute configurations were elucidated on the basis of an extensive spectroscopic analysis and quantum chemistry calculations. Compound 1 possesses a new carbon skeleton that might be derived from the protoilludane backbone. Compounds 1, 5, 8, and 9 show a certain degree of cytotoxicity to five human cancer cell lines. Compound 1 shows a mild inhibitory effect on nitric oxide production with an IC50 value of 31.4 µM. It is concluded that A. salicacola is rich in illudin derivatives with potential bioactivity prospects, which would make A. salicacola a good material of medicine and food homology.

Chemical constituents and their cytotoxicities from mushroom Tricholoma imbricatum.

Two undescribed triterpenes, tricholimbrins A and B, three undescribed steroids, tricholimbrins C‒E, one undescribed 4-chromanone derivative, along with 27 known compounds were isolated from fruiting bodies of the mushroom Tricholoma imbricatum. Tricholimbrins A and B are two polycyclic triterpenoids with a carbon degradation, while tricholimbrin C is a ring-rearranged steroid containing an aromatic moiety that might be derived from an ergosterol. Isocyathisterol, 3ß,15α-dihydroxyl-(22E,24R)-ergosta-5,8(14),22-trien-7-one, demethylincisterol A3, and volemolide showed cytotoxicities to six human cancer cell lines. 3ß-Hydroxyl-(22E,24R)-ergosta-5,8,22-trien-7,15-dione and 3ß-hydroxyl-(22E,24R)-ergosta-5,8,22-trien-7-one showed preferable cytotoxicities against HL-60 while chaxine C and volemolide showed preferable cytotoxicities against A-549, with IC50 values less than 10 µM.

Monoterpenoid indole alkaloids from the bark of Melodinus henryi.

Four new alkaloids, melodinines W1-W4 (1-4), together with twenty one known alkaloids (5-25) were isolated from Melodinus henryi. The structures with absolute configurations were elucidated by extensive MS and NMR spectroscopic methods, as well as the single crystal X-ray diffraction and ECD calculations. All compounds were evaluated for their cytotoxicities to five human cancer cell lines. Many compounds showed certain cytotoxicities to five human cancer cell lines with an IC50 range of 1.4-29.4 µM.