Increasing structure diversity of farnesylated chalcones by a fungal aromatic prenyltransferase.

Farnesylated chalcones were favored by researchers due to their different biological activities. However, only five naturally occurring farnesylated chalcones were described in the literature until now. Here, the farnesylation of six chalcones by the Aspergillus terreus aromatic prenyltransferase AtaPT was reported. Fourteen monofarnesylated chalcones (1F1-1F5, 2F1-2F3, 3F1, 3F2, 4F1, 4F2, 5F1, 6F1, and 6F2) and a difarnesylated product (2F3) were obtained, enriching the diversity of natural farnesylated chalcones significantly. Ten of them are C-farnesylated products, which complement O-farnesylated chalcones by chemical synthesis. Fourteen products have not been reported prior to this study. Nine of the produced compounds (1F2-1F5, 2F1-2F3, 5F1, and 6F1) exhibited inhibitory effect on α-glucosidase with IC50 values ranging from 24.08 ± 1.44 to 190.0 ± 0.28 µM. Among them, compounds 2F3 with IC50 value at 24.08 ± 1.44 µM and 1F4 with IC50 value at 30.09 ± 0.59 µM showed about 20 times stronger than the positive control acarbose with an IC50 at 536.87 ± 24.25 µM in α-glucosidase inhibitory assays.

Fungal Prenyltransferase AnaPT and Its F265 Mutants Catalyze the Dimethylallylation at the Nonaromatic Carbon of Phloretin.

Phloretin is widely found in fruit and shows various biological activities. Here, we demonstrate the dimethylallylation, geranylation, and farnesylation, particularly the first dimethylallylation at the nonaromatic carbon of phloretin (1) by the fungal prenyltransferase AnaPT and its mutants. F265 was identified as a key amino acid residue related to dimethylallylation at the nonaromatic carbon of phloretin. Mutants AnaPT_F265D, AnaPT_F265G, AnaPT_F265P, AnaPT_F265C, and AnaPT_F265Y were discovered to generally increase prenylation activity toward 1. AnaPT_F265G catalyzes the O-geranylation selectively at the C-2' hydroxyl group, which involves an intramolecular hydrogen bond with the carbonyl group of 1. Seven products, 1D5, 1D7-1D9, 1G2, 1G4, and 1F2, have not been reported prior to this study. Twelve compounds, 1D3-1D9, 1G1-1G3, and 1F1-1F2, exhibited potential inhibitory effects on α-glucosidase with IC50 values ranging from 11.45 ± 0.87 to 193.80 ± 6.52 µg/mL. Among them, 1G1 with an IC50 value of 11.45 ± 0.87 µg/mL was the most potential α-glucosidase inhibitor, which is about 30 times stronger than the positive control acarbose with an IC50 value of 346.63 ± 15.65 µg/mL.

Competitive coordination assembly of light-degradable gold nanocluster-intercalated metal organic frameworks for photoresponsive drug release.

On-demand controlled drug release holds great promise for cancer therapy. Light-degradable nanocarriers have gained increasing attention for designing controllable drug delivery systems owing to their spatiotemporally controllable properties. Herein, a highly luminescent and light-degradable nanocarrier is constructed by intercalating glutathione-capped gold nanoclusters (AuNCs) into zeolitic imidazolate framework-8 (ZIF-8) via competitive coordination assembly, named AuNC@ZIF-8, for light-triggered drug release. Glutathione-capped AuNCs and 2-methylimidazole (MIm) competitively coordinated with Zn2+ to form AuNC@ZIF-8 using a one step process in an aqueous solution. Specifically, the obtained AuNC@ZIF-8 has a high quantum yield of 52.96% and displays a distinctive property of photolysis. Competitive coordination interactions within AuNC@ZIF-8 were evidenced by X-ray diffraction and X-ray photoelectron spectroscopy, in which Zn2+ strongly coordinated with the N of MIm and weakly coordinated with the carboxyl/amino groups in the glutathione of AuNCs. Under light irradiation, the Au-S bond in AuNCs breaks, enhancing the coordination ability between carboxyl/amino groups and Zn2+. This collapses the crystal structure of AuNC@ZIF-8 and causes subsequent fluorescence quenching. Additionally, AuNC@ZIF-8 is successfully employed as a luminescent nanocarrier of anticancer drugs to form drug-AuNC@ZIF-8, in which three typical anticancer drugs are selected due to different coordination interactions. The obtained smart drug-AuNC@ZIF-8 can be effectively internalized into HeLa cells and degraded in response to blue light, with negligible dark cytotoxicity and high light cytotoxicity. This study highlights the crucial role of competitive coordination interactions in synthesizing functional materials with fluorescence efficiency and photolytic properties.

The Promiscuous Flavin-Dependent Monooxygenase PboD from Aspergillus ustus Increases the Structural Diversity of Hydroxylated Pyrroloindoline Diketopiperazines.

The potential of natural products as pharmaceutical and agricultural agents is based on their large structural diversity, resulting in part from modifications of the backbone structure by tailoring enzymes during biosynthesis. Flavin-dependent monooxygenases (FMOs), as one such group of enzymes, play an important role in the biosynthesis of diverse natural products, including cyclodipeptide (CDP) derivatives. The FMO PboD was shown to catalyze C-3 hydroxylation at the indole ring of cyclo-l-Trp-l-Leu in the biosynthesis of protubonines, accompanied by pyrrolidine ring formation. PboD substrate promiscuity was investigated in this study by testing its catalytic activity toward additional tryptophan-containing CDPs in vitro and biotransformation in Aspergillus nidulans transformants bearing a truncated protubonine gene cluster with pboD and two acetyltransferase genes. High acceptance of five CDPs was detected for PboD, especially of those with a second aromatic moiety. Isolation and structure elucidation of five pyrrolidine diketopiperazine products, with two new structures, proved the expected stereospecific hydroxylation and pyrrolidine ring formation. Determination of kinetic parameters revealed higher catalytic efficiency of PboD toward three CDPs consisting of aromatic amino acids than of its natural substrate cyclo-l-Trp-l-Leu. In the biotransformation experiments with the A. nidulans transformant, modest formation of hydroxylated and acetylated products was also detected.

Asperustins A-J: Austocystins with Immunosuppressive and Cytotoxic Activities from Aspergillus ustus NRRL 5856.

Ten new (1-10) and nine known (11-19) austocystins, along with four known anthraquinones (20-23), were isolated from the culture of Aspergillus ustus NRRL 5856 by bioactivity-guided fractionation. The structures of the new compounds were elucidated by spectroscopic data analysis, X-ray crystallographic study, the modified Mosher's method, [Rh2(OCOCF3)4]-induced ECD spectral analysis, and comparison of the experimental ECD spectra with those of the similar analogues. Compounds 1-8 represent the first examples of austocystins with a C-4' oxygenated substitution. The absolute configuration of 1″-hydroxy austocystin D (11) was determined by single-crystal X-ray diffraction and consideration of its biosynthetic origin. Compounds 5, 9, and 11 exhibited significant inhibitory effects against the proliferation of ConA-induced T cells with IC50 values of 1.1, 1.0, and 0.93 µM, respectively. Furthermore, these compounds suppressed the expression of IL-6 in a dose-dependent manner. Compounds 10-12 and 14 showed pronounced cytotoxicities against MCF-7 with IC50 values of 3.9, 1.3, 0.46, and 2.3 µM, respectively.

Biosynthesis of Epipyrone A Reveals a Highly Specific Membrane-Bound Fungal C-Glycosyltransferase for Pyrone Galactosylation.

Epipyrone A is a unique C-galactosylated 4-hydroxy-2-pyrone derivative with an antifungal potential from the fungus Epicoccum nigrum. We elucidated its biosynthesis via heterologous expression and characterized an unprecedented membrane-bound pyrone C-glycosyltransferase biochemically. Molecular docking and mutagenesis experiments suggested a possible mechanism for the heterocyclic C-glycosylation and the importance of a transmembrane helix for its catalysis. These results expand the repertoire of C-glycosyltransferases and provide new insights into the formation of C-glycosides in fungi.

Various interventions for cancer-related fatigue in patients with breast cancer: a systematic review and network meta-analysis.

Purpose: To investigate the effects of various intervention approaches on cancer-related fatigue (CRF) in patients with breast cancer. Method: Computer searches were conducted on PubMed, Embase, Cochrane Library, Web of Science, China National Knowledge Infrastructure (CNKI), China Science and Technology Journal Database (VIP), and Wanfang databases from their establishment to June 2023. Selection was made using inclusion and exclusion criteria, and 77 articles were included to compare the effects of 12 interventions on patients with breast cancer. Results: Seventy-seven studies with 12 various interventions were examined. The network findings indicated that cognitive behavioral therapy (CBT) (SMD, -1.56; 95%CI, -3.08~-0.04), Chinese traditional exercises (CTE) (SMD, -0.85; 95%CI, -1.34~-0.36), aerobic exercise (AE) (SMD, -0.77; 95%CI, -1.09~-0.45), multimodal exercise (ME) (SMD, -0.75; 95%CI, -1.26~-0.25), music interventions (MI) (SMD, -0.74; 95%CI, -1.45~-0.03), and yoga (YG) (SMD, -0.44; 95%CI, -0.83 to -0.06) can reduce CRF more than the control group (CG). For relaxation exercises (RE) (MD, -6.69; 95%CI, -9.81~-3.57), MI (MD, -5.45; 95%CI, -7.98~-2.92), AE (MD, -4.34; 95%CI, -5.90~-2.78), ME (MD, -3.47; 95%CI, -4.95~-1.99), YG (MD, -2.07; 95%CI, -3.56~-0.57), and mindfulness training (MD, -1.68; 95%CI, -2.91~-0.46), PSQI improvement was superior to CG. In addition, for CTE (MD, 11.39; 95%CI, 4.11-18.66), YG (MD, 11.28; 95%CI, 1.63-20.93), and AE (MD, 9.34; 95%CI, 0.26~18.42), Functional Assessment of Cancer Therapy-Breast improvement was superior to CG. Conclusion: Cognitive behavioral therapy (CBT) is the most effective measure for alleviating CRF in patients with breast cancer and Relaxation exercises (RE) is the most effective measure for improving sleep quality. In addition, Chinese traditional exercises (CTE) is the best measure for enhancing quality of life. Additional randomized controlled trials (RCTs) are expected to further investigate the efficacy and mechanisms of these interventions. Systematic review registration: https://www.crd.york.ac.uk/prospero/, identifier CRD42023471574.

Photocatalytic antibacterial agents based on inorganic semiconductor nanomaterials: a review.

Microbial contamination and antibiotic pollution have threatened public health and it is important to develop a rapid and safe sterilization strategy. Among various disinfection strategies, photocatalytic antibacterial methods have drawn increasing attention due to their efficient disinfection performances and environment-friendly properties. Although there are some reviews about bacterial disinfection, specific reviews on photocatalysis focused on inorganic semiconductor nanomaterials are rarely reported. Herein, we present a systematic summary of recent disinfection developments based on inorganic nanomaterials (including metal oxides, sulfides, phosphides, carbon materials, and corresponding heterostructures) over the past five years. Moreover, key factors and challenges for inorganic nanomaterial-based photocatalytic disinfection are outlined, which holds great potential for future photocatalytic antibacterial applications.

Microglia Promote Inhibitory Synapse Phagocytosis in the Spinal Cord Dorsal Horn and Modulate Pain-Like Behaviors in a Murine Cancer-Induced Bone Pain Model.

BACKGROUND: The microglial activation has been implicated in cancer-induced bone pain. Recent studies have revealed that microglia mediate synaptic pruning in the central nervous system, where the cluster of differentiation 47-signal regulatory protein α (CD47-SIRPα) axis creates a "don't eat me" signal and elicits an antiphagocytic effect to protect synapses against elimination. To date, the synaptic phagocytosis in microglia has never been investigated in the murine cancer-induced bone pain model. The present experiments sought to explore whether microglia phagocytize synapses in mice with bone cancer pain as well as the possible mechanisms. METHODS: Male C3H/HeN mice were used to induce bone cancer pain. Minocycline and S-ketamine were injected into D14. The number of spontaneous flinches (NSF) and paw withdrawal mechanical thresholds (PWMT) were measured on D0, D4, D7, D10, D14, D21, and D28. Hematoxylin and eosin staining presented bone lesions. Western blotting examined the Gephyrin, CD47, and SIRPα expression. Flow cytometry evaluated the proportion of SIRPα+ cells in the spine. Immunofluorescence and 3-dimensional reconstruction showed the Gephyrin puncta inside microglial lysosomes. RESULTS: Mice embedded with tumor cells induced persistent spontaneous pain and mechanical hyperalgesia. Hematoxylin and eosin staining revealed bone destruction and tumor infiltration in marrow cavities. Microglia underwent a responsive and proliferative burst (t = -16.831, P < .001). Western blotting manifested lowered Gephyrin expression in the tumor group (D4, D7, D10, D14, D21, and D28: P < .001). Immunofluorescence and 3-dimensional reconstruction showed larger volumes of Gephyrin puncta inside microglial lysosomes (t = -23.273, P < .001; t = -27.997, P < .001). Treatment with minocycline or S-ketamine exhibited pain relief and antiphagocytic effects (t = -6.191, P < .001, t = -7.083, P < .001; t = -20.767, P < .001, t = -17.080, P < .001; t = 11.789, P < .001, t = 16.777, P < .001; t = 8.868, P < .001, t = 21.319, P < .001). Last but not least, the levels of CD47 and SIRPα proteins were downregulated (D10: P = .004, D14, D21, and D28: P < .001; D10, D14, D21, and D28: P < .001). Flow cytometry and immunofluorescence substantiated reduced microglial SIRPα (t = 11.311, P < .001; t = 12.189, P < .001). CONCLUSIONS: Microglia-mediated GABAergic synapse pruning in the spinal cord dorsal horn in bone cancer pain mice, which might be associated with the declined CD47-SIRPα signal. Our research uncovered an innovative mechanism that highlighted microglia-mediated synaptic phagocytosis in a murine cancer-induced bone pain model.

Hollow structured CdS@ZnIn2S4 Z-scheme heterojunction for bifunctional photocatalytic hydrogen evolution and selective benzylamine oxidation.

Photocatalytic hydrogen evolution (PHE) is frequently constrained by inadequate light utilization and the rapid combination rate of the photogenerated electron-hole pairs. Additionally, conventional PHE processes are often facilitated by the addition of sacrificial reagents to consume photo-induced holes, which makes this approach economically unfavorable. Herein, we designed a spatially separated bifunctional cocatalyst decorated Z-scheme heterojunction of hollow structured CdS (HCdS) @ZnIn2S4 (ZIS), which was prepared by a sacrificial hard template method followed by photo-deposition. Consequently, PdOx@HCdS@ZIS@Pt exhibited efficient PHE (86.38 mmol·g-1·h-1) and benzylamine (BA) oxidation coupling (164.75 mmol·g-1·h-1) with high selectivity (97.34 %). The unique hollow core-shelled morphology and bifunctional cocatalyst loading in this work hold great potential for the design and synthesis of bifunctional Z-scheme photocatalysts.

Four new steroidal glycosides from Lilium lancifolium Thunb. and their antitumor activity.

Four new steroidal glycosides (1-4), including two steroidal saponins named lililancifoloside B and C (1-2), one pregnane glycoside named lililancifoloside D (3), and one C22-steroidal lactone glycoside named lililancifoloside E (4), together with five known ones (5-9), were isolated from the bulbs of Lilium lancifolium Thunb. By using spectroscopic analysis, including 1D, 2D NMR, and HR-ESI-MS, the structures of 1-4 were elucidated. All isolates were tested for their cytotoxic potential against the MCF-7, MDA-MB-231, HepG2, and A549 cell lines. Compound 6 distinguished out among them, IC50 values of 3.31, 5.23, 1.78, and 1.49 µM against the four cell lines, respectively. Other compounds such as compound 3, 5, and 9 have also shown specific cytotoxic activity. Next, studies showed that compound 6 might cause HepG2 cells to undergo a cell cycle arrest during the G2/M phase and apoptosis.

Genome-wide characterization of microsatellites in cobia Rachycentron canadum (Linnaeus, 1766): Survey and analysis of their abundance and diversity.

The cobia Rachycentron canadum, mainly distributed in the warm waters of tropical and subtropical regions around the world, remains a fish of considerable economic importance. Detailed diversity and the number of microsatellite sequences in the cobia genome are still unintelligible. The primary aim of this work was to identify and quantify the miscellaneous SSR sequences in the cobia genome. More than 280,000 sequences were sequenced and screened using next-generation sequencing technology and microsatellite identification. Perfect mononucleotide repeats, dinucleotide microsatellites, and trinucleotide microsatellites contain (A)10 /(T)10 , (AC)6 /(TG)6 , and (AAT)5-32 as the largest number of motifs in each type of microsatellite, respectively. The tetranucleotide and pentanucleotide microsatellites (TTM and PTM) consist of the largest number of motifs of both (ATCT)5-32 and (TCAT)5-31 in TTMs, and (CTCTC)5-9 in PTMs, whereas the hexanucleotide microsatellites are rarely observed in the cobia genome. All c. 38000 sequences of composite microsatellites are extremely diverse, including compound (11.71%), interrupted compound (71.77%), complex (0.45%), and interrupted complex (16.07%). In this study, we developed a convenient and useful recording system for writing down and categorizing diverse composite microsatellite types. This system will provide great support for exploring repeat origins, evolutionary mechanisms, and the application of polymorphic microsatellites.

P450 in C-C coupling of cyclodipeptides with nucleobases.

Bacterial cytochrome P450 enzymes catalyze various and often intriguing tailoring reactions during the biosynthesis of natural products. In contrast to the majority of membrane-bound P450 enzymes from eukaryotes, bacterial P450 enzymes are soluble proteins and therefore represent excellent candidates for in vitro biochemical investigations. In particular, cyclodipeptide synthase-associated cytochrome P450 enzymes have recently gained attention due to the broad spectrum of reactions they catalyze, i.e. hydroxylation, aromatization, intramolecular C-C bond formation, dimerization, and nucleobase addition. The latter reaction has been described during the biosynthesis of guanitrypmycins, guatrypmethines and guatyromycines in various Streptomyces strains, where the nucleobases guanine and hypoxanthine are coupled to cyclodipeptides via C-C, C-N, and C-O bonds. In this chapter, we provide an overview of cytochrome P450 enzymes involved in the C-C coupling of cyclodipeptides with nucleobases and describe the protocols used for the successful characterization of these enzymes in our laboratory. The procedure includes cloning of the respective genes into expression vectors and subsequent overproduction of the corresponding proteins in E. coli as well as heterologous expression in Streptomyces. We describe the purification and in vitro biochemical characterization of the enzymes and protocols to isolate the produced compounds for structure elucidation.

Causality of occupational exposure on rheumatoid arthritis and ankylosing spondylitis: a two-sample mendelian randomization study.

Objective: This study aimed to explore the potential causal link between three specific types of occupational exposure on rheumatoid arthritis (RA) and ankylosing spondylitis (AS). Method: A Two-sample Mendelian randomization (TSMR) analysis, comprising univariate MR (UVMR) and multivariate MR (MVMR) analyses, was performed to investigate the potential causal association between three types of occupational exposures, jobs involving mainly walking or standing (JWS), jobs involving heavy manual or physical work (JMP), and jobs involving shift work(JSW) on RA and AS. Genetic variants for genome-wide association studies (GWAS) of occupational exposure and AS were obtained from the UK Biobank. GWAS summary data for RA were obtained from FinnGen Biobank analysis. For UVMR, six methods of Inverse Variance Weighted (IVW), MR-Egger, Weighted Mode, Weighted Median, Simple Mode, MR pleiotropy residual sum, and outlier (MR-PRESSO) were used for the analysis. The MVMR was analyzed using the IVW model as well as the MR-Egger model. Results: The UVMR suggested no causal relationship between the three occupational exposure and RA [IVW: P=0.59,0.21,0.63] or AS [IVW: P=0.43,0.57,0.04], as did the bidirectional MR [IVW: P=0.73,0.70,0.16], [IVW: P=0.65,0.68,0.74]. Although unadjusted MVMR suggested a causal relationship between JMP and AS [IVW: OR = 1.01, 95% CI = 1.00- 1.02, p = 0.02], the adjusted MVMR denied this relationship and concluded that there was no causal relationship between the other occupational exposure and either RA or AS. Conclusion: Our MR analysis did not establish a direct causal relationship between certain occupational exposures and either RA or AS.

Prenylation of dimeric cyclo-L-Trp-L-Trp by the promiscuous cyclo-L-Trp-L-Ala prenyltransferase EchPT1.

Prenyltransferases (PTs) from the dimethylallyl tryptophan synthase (DMATS) superfamily are known as efficient biocatalysts and mainly catalyze regioselective Friedel-Crafts alkylation of tryptophan and tryptophan-containing cyclodipeptides (CDPs). They can also use other unnatural aromatic compounds as substrates and play therefore a pivotal role in increasing structural diversity and biological activities of a broad range of natural and unnatural products. In recent years, several prenylated dimeric CDPs have been identified with wide range of bioactivities. In this study, we demonstrate the production of prenylated dimeric CDPs by chemoenzymatic synthesis with a known promiscuous enzyme EchPT1, which uses cyclo-L-Trp-L-Ala as natural substrate for reverse C2-prenylation. High product yields were achieved with EchPT1 for C3-N1' and C3-C3' linked dimers of cyclo-L-Trp-L-Trp. Isolation and structural elucidation confirmed the product structures to be reversely C19/C19'-mono- and diprenylated cyclo-L-Trp-L-Trp dimers. Our study provides an additional example for increasing structural diversity by prenylation of complex substrates with known biosynthetic enzymes. KEY POINTS: • Chemoenzymatic synthesis of prenylated cyclo-L-Trp-L-Trp dimers • Same prenylation pattern and position for cyclodipeptides and their dimers. • Indole prenyltransferases such as EchPT1 can be widely used as biocatalysts.

Biosynthesis of p-Terphenyls in Aspergillus ustus Implies Enzymatic Reductive Dehydration and Spontaneous Dibenzofuran Formation.

p-Terphenyls contain a central benzene ring substituted with two phenyl residues at its para positions. Heterologous expression of a biosynthetic gene cluster from Aspergillus ustus led to the formation of four new p-terphenyl derivatives. Gene deletion experiments proved the formation and reductive dehydration of the terphenylquinone atromentin, followed by O-methylation and prenylation. Spontaneous dibenzofuran formation led to the final products. These results provide new insights into the biosynthesis of p-terphenyls in fungi and dibenzofuran formation in the biosynthesis of numerous natural products.

Fabrication of Al/AlOx/Al junctions with high uniformity and stability on sapphire substrates.

Tantalum and aluminum on sapphire are widely used platforms for qubits of long coherent time. As quantum chips scale up, the number of Josephson junctions on sapphire increases. Thus, both the uniformity and stability of the junctions are crucial to quantum devices, such as scalable superconducting quantum computer circuit, and quantum-limited amplifiers. By optimizing the fabrication process, especially, the conductive layer during the electron beam lithography process, Al/AlOx/Al junctions of sizes ranging from 0.0169 to 0.04 µm2 on sapphire substrates were prepared. The relative standard deviation of room temperature resistances (RN) - [Formula: see text] of these junctions is better than 1.7% on 15 mm × 15 mm chips, and better than 2.66% on 2 inch wafers, which is the highest uniformity on sapphire substrates has been reported. The junctions are robust and stable in resistances as temperature changes. The resistances increase by the ratio of 9.73% relative to RN as the temperature ramp down to 4 K, and restore their initial values in the reverse process as the temperature ramps back to room temperature. After being stored in a nitrogen cabinet for 100 days, the resistance of the junctions changed by1.16% on average. The demonstration of uniform and stable Josephson junctions in large area paves the way for the fabrication of superconducting chip of hundreds of qubits on sapphire substrates.

Colletotriauxins A-D, New Plant Growth Inhibitors from the Phytopathogenic Fungus Colletotrichum gloeosporioides.

Four undescribed plant growth inhibitory indole derivatives, colletotriauxins A-D (1-4), along with two known analogues indole-3-acetic acid (IAA) (5) and its amide indole-3-acetamide (6), were isolated from the phytopathogenic fungus Colletotrichum gloeosporioides NRRL 45420. Their structures were elucidated by NMR and MS analyses. 1 and 2 are rhamnosides of indole-3-ethanol (tryptophol) and its methylated derivative, respectively. In the structures of 3 and 4, the two terminal hydroxyl groups of hexitol and pentane-1,2,3,4,5-pentol are connected with indole-3-(2-methyl)-acetyl and acetyl moieties. Compounds 1-6 inhibit Lepidium sativum seedling growth. The inhibition activities of colletotriauxins for stem growth were even stronger than IAA, with compounds 3 and 4 as the most active ones. These results suggested that colletotriauxins could serve as potential candidates as herbicides.

A Flavin-Dependent Oxygenase Catalyzes Hydroxylation and Simultaneous Pyrrolidine Ring Formation in Protubonine Biosynthesis in Aspergillus ustus.

The hydroxylated and diacetylated cyclo-l-Trp-l-Leu derivative (-)-protubonine B was isolated from a culture of Aspergillus ustus 3.3904. Genome mining led to the identification of a putative biosynthetic gene cluster coding for a bimodular nonribosomal peptide synthetase, a flavin-dependent monooxygenase, and two acetyltransferases. Heterologous expression of the pbo cluster in Aspergillus nidulans showed that it is responsible for the formation of the isolated metabolite. Gene deletion experiments and structural elucidation of the isolated intermediates confirmed the biosynthetic steps. In vitro experiments with the recombinant protein proved that the flavin-dependent oxygenase is responsible for stereospecific hydroxylation at the indole ring accompanied by pyrrolidine ring formation.

A basidomycetous hydroxynaphthalene-prenylating enzyme exhibits promiscuity toward prenyl donors.

The fungal prenyltransferase ShPT from Stereum hirsutum was believed to prenylate 4-hydroxybenzyl alcohol and thereby be involved in the vibralactone biosynthesis. In this study, we demonstrate that hydroxynaphthalenes instead of benzyl alcohol or aldehyde were accepted by ShPT for regular C-prenylation in the presence of both dimethylallyl and geranyl diphosphate. Although the natural substrate of ShPT remains unknown, our results provide one additional prenyltransferase from basidiomycetes, which are less studied, in comparison to those from other sources. Furthermore, this study expands the chemical toolbox for regioselective production of prenylated naphthalene derivatives. KEY POINTS: •Basidiomycetous prenyltransferase •Biochemical characterization •A DMATS prenyltransferase prenylating hydroxynaphthalene derivatives.