Sorbremnoids A and B: NLRP3 Inflammasome Inhibitors Discovered from Spatially Restricted Crosstalk of Biosynthetic Pathways.

Crosstalk-oriented chemical evolution of natural products (NPs) is an efficacious strategy for generating novel skeletons through coupling reactions between NP fragments. In this study, two NOD-like receptor protein 3 (NLRP3) inflammasome inhibitors, sorbremnoids A and B (1 and 2), with unprecedented chemical architectures were identified from a fungus Penicillium citrinum. Compounds 1 and 2 exemplify rare instances of hybrid NPs formed via a major facilitator superfamily (MFS)-like enzyme by coupling reactive intermediates from two separate biosynthetic gene clusters (BGCs), pcisor and pci56. Both sorbremnoids A and B are NLRP3 inflammasome inhibitors. Sorbremnoid A demonstrated strong inhibition of IL-1ß by directly binding to the NLRP3 protein, inhibiting the assembly and activation of the NLRP3 inflammasome in vitro, with potential application in diabetic refractory wound healing through the suppression of excessive inflammatory responses. This research will inspire the development of anti-NLRP3 inflammasome agents as lead treatments and enhance knowledge pertaining to NPs derived from biosynthetic crosstalk.

Cyclo-diphenylalanine production in Aspergillus nidulans through stepwise metabolic engineering.

Cyclo-diphenylalanine (cFF) is a symmetrical aromatic diketopiperazine (DKP) found wide-spread in microbes, plants, and resulting food products. As different bioactivities continue being discovered and relevant food and pharmaceutical applications gradually emerge for cFF, there is a growing need for establishing convenient and efficient methods to access this type of compound. Here, we present a robust cFF production system which entailed stepwise engineering of the filamentous fungal strain Aspergillus nidulans A1145 as a heterologous expression host. We first established a preliminary cFF producing strain by introducing the heterologous nonribosomal peptide synthetase (NRPS) gene penP1 to A. nidulans A1145. Key metabolic pathways involving shikimate and aromatic amino acid biosynthetic support were then engineered through a combination of gene deletions of competitive pathway steps, over-expressing feedback-insensitive enzymes in phenylalanine biosynthesis, and introducing a phosphoketolase-based pathway, which diverted glycolytic flux toward the formation of erythrose 4-phosphate (E4P). Through the stepwise engineering of A. nidulans A1145 outlined above, involving both heterologous pathway addition and native pathway metabolic engineering, we were able to produce cFF with titers reaching 611 mg/L in shake flask culture and 2.5 g/L in bench-scale fed-batch bioreactor culture. Our study establishes a production platform for cFF biosynthesis and successfully demonstrates engineering of phenylalanine derived diketopiperazines in a filamentous fungal host.

Evolution-Based Discovery of Polyketide Acylated Valine from a Cytochalasin-Like Gene Cluster in Simplicillium lamelliciola HDN13430.

Utilizing a gene evolution-oriented approach for gene cluster mining, a cryptic cytochalasin-like gene cluster (sla) in Antarctic-derived Simplicillium lamelliciola HDN13430 was identified. Compared with the canonical cytochalasin biosynthetic gene clusters (BGCs), the sla gene cluster lacks the key α,ß-hydrolase gene. Heterologous expression of the sla gene cluster led to the discovery of a new compound, slamysin (1), characterized by an N-acylated amino acid structure and demonstrating weak anti-Bacillus cereus activity. These findings underscore the potential of genetic evolution in uncovering novel compounds and indicating specific adaptive evolution within specialized habitats.

Phenylhydrazone Alkaloids from the Deep-Sea Cold Seep Derived Fungus Talaromyces amestolkiae HDN21-0307.

Alkaloids with a phenylhydrazone architecture are rarely found in nature. Four unusual phenylhydrazone alkaloids named talarohydrazones A-D (1-4) were isolated from the deep-sea cold seep derived fungus Talaromyces amestolkiae HDN21-0307 using the one strain-many compounds (OSMAC) approach and MS/MS-based molecular networking (MN) combined with network annotation propagation (NAP) and the unsupervised substructure annotation method MS2LDA. Their structures were elucidated by spectroscopic data analysis, single-crystal X-ray diffraction, and quantum chemical calculations. Talarohydrazone A (1) possessed an unusual skeleton combining 2,4-pyridinedione and phenylhydrazone. Talarohydrazone B (2) represents the first natural phenylhydrazone-bearing azadophilone. Bioactivity evaluation revealed that compound 1 exhibited cytotoxic activity against NCI-H446 cells with an IC50 value of 4.1 µM. In addition, compound 1 displayed weak antibacterial activity toward Staphylococcus aureus with an MIC value of 32 µg/mL.

Impacts of irrigation with treated livestock wastewater on the accumulation characteristic of ARGs in the farmland soil: a case study in Hohhot, China.

Irrigation with treated livestock wastewater (TWW) is a promising strategy for reusing resources. However, TWW irrigation might introduce antibiotic resistant genes (ARGs) into the soil, posing environmental risks associated with antibiotic resistance. This study focuses on investigating the influence of irrigation amounts and duration on the fate of ARGs and identifies key factors driving their changes. The results showed that there were 13 ARGs in TWW, while only 5 ARGs were detected in irrigated soil. That is some introduced ARGs from TWW could not persistently exist in the soil. After 1-year irrigation, an increase in irrigation amount from 0.016 t/m2 to 0.048 t/m2 significantly enhanced the abundance of tetC by 29.81%, while ermB and sul2 decreased by 45.37% and 76.47%, respectively (p < 0.01). After 2-year irrigation, the abundance of tetC, ermB, ermF, dfrA1, and total ARGs significantly increased (p < 0.05) when the irrigation amount increased. The abundances of ARGs after 2-year irrigation were found to be 2.5-34.4 times higher than 1 year. Obviously, the irrigation years intensified the positive correlation between ARGs abundance and irrigation amount. TetC and ermF were the dominant genes resulting in the accumulation of ARGs. TWW irrigation increased the content of organic matter and total nitrogen in the soil, which affected microbial community structure. The changes of the potential host were the determining factors driving the ARGs abundance. Our study demonstrated that continuous TWW irrigation for 2 years led to a substantial accumulation of ARGs in soil.

Discovery, Total Synthesis, and Anti-Inflammatory Evaluation of Naturally Occurring Naphthopyrone-Macrolide Hybrids as Potent NLRP3 Inflammasome Inhibitors.

Numerous clinical disorders have been linked to the etiology of dysregulated NLRP3 (NACHT, LRR, and PYD domain-containing protein 3) inflammasome activation. Despite its potential as a pharmacological target, modulation of NLRP3 activity remains challenging. Only a sparse number of compounds have been reported that can modulate NLRP3 and none of them have been developed into a commercially available drug. In this research, we identified three potent NLRP3 inflammasome inhibitors, gymnoasins A-C (1-3), with unprecedented pentacyclic scaffolds, from an Antarctic fungus Pseudogymnoascus sp. HDN17-895, which represent the first naturally occurring naphthopyrone-macrolide hybrids. Additionally, biomimetic synthesis of gymnoasin A (1) was also achieved validating the chemical structure and affording ample amounts of material for exhaustive bioactivity assessments. Biological assays indicated that 1 could significantly inhibited in vitro NLRP3 inflammasome activation and in vivo pro-inflammatory cytokine IL-1ß release, representing a valuable new lead compound for the development of novel therapeutics with the potential to inhibit the NLRP3 inflammasome.

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.

Leveraging machine learning for prediction of antibiotic resistance genes post thermal hydrolysis-anaerobic digestion in dairy waste.

Anaerobic digestion holds promise as a method for removing antibiotic resistance genes (ARGs) from dairy waste. However, accurately predicting the efficiency of ARG removal remains a challenge. This study introduces a novel appproach utilizing machine learning to forecast changes in ARG abundances following thermal hydrolysis-anaerobic digestion (TH-AD) treatment. Through network analysis and redundancy analyses, key determinants of affect ARG fluctuations were identified, facilitating the development of machine learning models capable of accurately predicting ARG changes during TH-AD processes. The decision tree model demonstrated impressive predictive power, achieving an impessive R2 value of 87% against validation data. Feature analysis revealed that the genes intI2 and intI1 had a critical impact on the absolute abundance of ARGs. The predictive model developed in this study offers valuable insights for improving operational and managerial practices in dairy waste treatment facilities, with the ultimate goal of mitigating the spread of antibiotic resistance.

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.

Multifunctional Cross-Linking Composite Binder Enables the Stable Performance of Si-Based Anodes for High-Energy-Density Lithium-Ion Batteries.

The intrinsic volumetric stress during cycling is the main obstacle for developing Si-based materials as high-energy-density lithium-ion battery anodes. Elastic binders have been demonstrated as an efficient approach to alleviate the stress of Si. Herein, we design a tough 3D hard/soft polymeric network (LPTS) using lithiated poly(acrylic acid), silk sericin, and highly branched tannic acid. Covalent cross-linking provides a robust mechanical strength to endure the large stress. The formed multiple hydrogen bonds with bonding energies between 3.46 and 25 kcal mol-1 can effectively dissipate the stress through sequential hydrogen bond disassociation. The multifunctional LPTS binder maintains the integrity of the Si-based electrodes during repeated discharging/charging. Additionally, Li+ can be transferred via a Li-conducting group (-COOLi), thereby enhancing the ionic conductivity of electrodes. Consequently, the Si/LPTS electrode exhibits an improved initial Coulombic efficiency and excellent durability over 400 cycles. Meanwhile, this binder is also suitable for Si-C anodes, enabling stable cycling at a high areal capacity >3.6 mAh cm-2 and delivering 72.2% capacity retention for the LFP||Si-C/LPTS full cell after 200 cycles. This study provides insight into developing efficient Si-based binders that are facile and low-cost for next-generation high-energy-density systems.

Genome Mining Reveals a UbiA-Type Prenyltransferase Access to Farnesylation of Diketopiperazines.

UbiA-type prenyltransferases (PTases) are significant enzymes that lead to structurally diverse meroterpenoids. Herein, we report the identification and characterization of an undescribed UbiA-type PTase, FtaB, that is responsible for the farnesylation of indole-containing diketopiperazines (DKPs) through genome mining. Heterologous expression of the fta gene cluster and non-native pathways result in the production of a series of new C2-farnesylated DKPs. This study broadens the reaction scope of UbiA-type PTases and expands the chemical diversity of meroterpenoids.

Antibacterial p-terphenyl and α­pyrone derivates isolated from the marine-derived actinomycete Nocardiopsis sp. HDN154086.

Assisted by OSMAC strategy, one new p-terphenyl and two new α­pyrone derivates, namely nocarterphenyl I (1) and nocardiopyrone D-E (2-3), were obtained and characterized from the marine sediment-derived actinomycete Nocardiopsis sp. HDN154086. The structures of these compounds were determined on the basis of MS, NMR spectroscopic data and single-crystal X-ray diffraction. Compound 1 with a rare 2,2'-bithiazole structure among natural products showed promising activity against five bacteria with MIC values ranging from 0.8 to 1.6 µM and 3 exhibited notable antibacterial activity against MRSA compared the positive control ciprofloxacin.

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.