Heterologous production of bioactive xenoacremone analogs in Aspergillus nidulans / 中国天然药物

Tyrosine-decahydrofluorene derivatives are a class of hybrid compounds that integrate the properties of polyketides and nonribosomal peptides. These compounds feature a [6.5.6] tricarbocyclic core and a para-cyclophane ether moiety in their structures and exhibit anti-tumor and anti-microbial activities. In this study, we constructed the biosynthetic pathway of xenoacremones from Xenoacremonium sinensis ML-31 in the Aspergillus nidulans host, resulting in the identification of four novel tyrosine-decahydrofluorene analogs, xenoacremones I-L (1-4), along with two known analogs, xenoacremones A and B. Remarkably, compounds 3 and 4 contained a 12-membered para-cyclophane ring system, which is unprecedented among tyrosine-decahydrofluorene analogs in X. sinensis. The successful reconstruction of the biosynthetic pathway and the discovery of novel analogs demonstrate the utility of heterologous expression strategy for the generation of structurally diverse natural products with potential biological activities.

An epipolythiodioxopiperazine alkaloid and diversified aromatic polyketides with cytotoxicity from the Beibu Gulf coral-derived fungus Emericella nidulans GXIMD 02509 / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Marine microorganisms, especially marine fungi, have historically proven their value as a prolific source for structurally novel and pharmacologically active secondary metabolites (Deshmukh et al., 2018; Carroll et al., 2022). The corals constitute a dominant part of reefs with the highest biodiversity, and harbor highly diverse and abundant microbial symbionts in their tissue, skeleton, and mucus layer, with species-specific core members that are spatially partitioned across coral microhabitats (Wang WQ et al., 2022). The coral-associated fungi were very recently found to be vital producers of structurally diverse compounds, terpenes, alkaloids, peptides, aromatics, lactones, and steroids. They demonstrate a wide range of bioactivity such as anticancer, antimicrobial, and antifouling activity (Chen et al., 2022). The genetically powerful genus Emericella (Ascomycota), which has marine and terrestrial sources, includes over 30 species and is distributed worldwide. It is considered a rich source of diverse secondary metabolites with antimicrobial activity or cytotoxicity (Alburae et al., 2020). Notably, Emericella nidulans, the sexual state of a classic biosynthetic strain Aspergillus nidulans, was recently reported as an important source of highly methylated polyketides (Li et al., 2019) and isoindolone-containing meroterpenoids (Zhou et al., 2016) with unusual skeletons.

Screening of marine resistant strain based on PKS and NRPS genes and the activity of its metabolites / 生物工程学报

Based on polyketide syntheses gene (PKS) and non-ribosomal peptide synthetases gene (NRPS), one strain with high anti-pathogenic activity was screened from 77 strains isolated from Arctic marine sediments and identified. By optimizing the composition of culture medium and fermentation conditions, the production of this strain's active metabolites was improved and the main metabolites were identified by HRMS, 1H NMR and 13C NMR. The antibacterial spectrum of the main metabolites and the effect of the metabolites on cucumber Fusarium wilt were also determined. The results showed that the strain was Bacillus velezensis and it showed growth promoting effect on plants. When the strain was cultured in 5 g/L maltose, 10 g/L tryptone, 10 g/L sodium chloride, at 30 ℃, 150 r/min for 60 h, the diameter of the inhibition zone increased from (16.23±0.42) to (24.42±0.57) mm. The metabolites of this strain mainly contain macrolide compound macrolactin A, which has antagonistic effect on a variety of pathogenic bacteria and fungi. Cucumber seedling experiments showed that the metabolites of this strain had a protective effect on cucumber Fusarium wilt, and showed a good potential for development and application as a biocontrol agent.

Secondary metabolites of endophyte fungi Xylaria sp. from Coptis chinensis / 中国中药杂志

Two new polyketides, lasobutone A(1) and lasobutone B(2), along with three known compounds, guignardianone C(3), guignardic acid(4), and 4-hydroxy-17R-methylincisterol(5), were isolated from the endophytic fungi Xylaria sp. by silica gel, MCI, and preparative HPLC, which was separated from the Chinese medicinal material Coptis chinensis and cultivated through solid fermentation with rice. Their structures were elucidated on the basis of spectroscopic methods, such as MS, NMR, IR, UV, and ECD. Compounds 2 and 4 showed inhibitory activities against the nitric oxide(NO) production in the LPS-induced macrophage RAW264.7 with IC_(50) values of 58.7 and 42.5 μmol·L~(-1) respectively, while compound 5 exhibited cytotoxic activities against HT-29 with IC_(50) value of 14.3 μmol·L~(-1).

A new polyketide of endophytic fungi Aspergillus sp. ZJ-58 from Coptis chinensis / 中国中药杂志

A new polyketide, coptaspin A(1), along with two known compounds 4-acetyl-3,4-dihydro-6,8-dihydroxy-3-methoxy-5-methylisocoumarin(2), and cytochalasin Z_(12)(3), was isolated from the endophytic fungi Aspergillus sp. ZJ-58, which was isolated from the genuine medicinal plant Coptis chinensis in Chongqing after solid-state fermentation on rice and silica gel, MCI, and HPLC-based separation. Their structures were elucidated by MS, NMR, IR, UV, and ECD. The newly isolated compound 1 showed moderate inhibitory activities against LPS-induced NO production in RAW264.7 macrophages with the IC_(50) value of 58.7 μmol·L~(-1), suggesting its potential anti-inflammatory activity.

The recent advances in developing gene editing and expression tools and the synthesis of natural products in Yarrowia lipolytica / 生物工程学报

Yarrowia lipolytica, as an important oleaginous yeast, has been widely used in metabolic engineering. Y. lipolytica is considered as an ideal host for the production of natural products such as terpenes, polyketides and flavonoids, due to its ability to utilize a variety of hydrophobic substrates, high stress tolerance to acid and salt, high flux of tricarboxylic acid cycle and the ability in providing abundant the common precursor acetyl-CoA. Recently, more and more tools for genetic editing, gene expression and regulation has been developed in Y. lipolytica, which facilitate the metabolic engineering of Y. lipolytica for bio-manufacturing. In this review, we summarized the recent progresses in developing gene expression and natural product synthesis in Y. lipolytica, and also discussed the challenges and possible solutions in heterologous synthesis of natural products in this yeast.

Three new polyketides from vasR2 gene over-expressed mutant strain of Verrucosispora sp. NS0172 / 中国天然药物

Over-expression of the pathway specific positive regulator gene is an effective way to activate silent gene cluster. In the curret study, the SARP family regulatory gene, vasR2, was over-expressed in strain Verrucosispora sp. NS0172 and the cryptic gene cluster responsible for the biosynthesis of pentaketide ansamycin was partially activated. Two tetraketides (1 and 2) and a triketide (3) ansamycins, together with five known compounds (4-8), were isolated and elucidated from strain NS0172OEvasR2. Their NMR data were completely assigned by analysis of their HR-ESI-MS and

Novel angucycline/angucyclinone family of natural products discovered between 2010 and 2020 / 生物工程学报

Angucyclines/angucyclinones are a large group of polycyclic aromatic polyketides and their producers are widely distributed in nature. This family of natural products attracts great attention because of their diverse biological activities and unique chemical structures. With the development of synthetic biology and the exploitation of the actinomycetes from previously unexplored environments, angucyclines/angucyclinones-like natural products with new skeletons were continuously discovered, thus enriching the structural diversity of this family. In this review we summarize the new angucyclines/angucyclinones analogues discovered in the last decade (2010-2020) by using different strategies, such as changing cultivation conditions, genetic modification, genome mining, bioactivity-guided compound isolation, and fermentation of actinomycetes from underexplored environments. We also discuss the role of synthetic biology in the discovery and development of new compounds of the angucycline/angucyclinone family.

Research progress on natural products of fungi of Diaporthe sp / 中国中药杂志

Diaporthe sp. fungi is one of the important sources of active natural products. Polyketides, alkaloids, terpenes, anthraquinones and other types of novel metabolic products are found from this genus, and many of them have significant anti-tumor, antibacterial, anti-hyperlipidemia, inhibition of pulmonary fibrosis, antioxidant and other biological activities. This paper reviewed source, structure and biological activity of natural products from Diaporthe sp. in the past two decades, and provided a reference for in-depth study of natural product of this genus fungus and innovative drug development.

A new polyketide from marine-derived Streptomyces sp.MDW-06 / 中国中药杂志

To isolate and identify secondary metabolites of marine-derived Streptomyces sp.MDW-06,the isolations and purifications of compounds were performed by means of column chromatography over silica gel. And their structures were elucidated through the spectroscopic analysis of MS,NMR and specific rotations. The bioactivities were assayed by paper diffusion and DPPH method. From the fermentation broth of marine-derived Streptomyces sp.MDW-06,five compounds( 1-5) were isolated and identified as streptopentanoic acid( 1),germicidin A( 2),germicidin B( 3),isogermicidin A( 4),isogermicidin B( 5) and oxohygrolidin( 6),respectively. Compound 1 is a new compound. Compound 1 shows DPPH radical scavenging activity with 36. 4% at 100 mg·L~(-1).

Peptides and polyketides isolated from the marine sponge-derived fungus Aspergillus terreus SCSIO 41008 / 中国天然药物

Two new isomeric modified tripeptides, aspergillamides C and D (compounds 1 and 2), together with fifteen known compounds (compounds 3-17), were obtained from the marine sponge-derived fungus Aspergillus terreus SCSIO 41008. The structures of the new compounds, including absolute configurations, were determined by extensive analyses of spectroscopic data (NMR, MS, UV, and IR) and comparisons between the calculated and experimental electronic circular dichroism (ECD) spectra. Butyrolactone I (compound 11) exhibited strong inhibitory effects against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with the IC being 5.11 ± 0.53 μmol·L, and acted as a noncompetitive inhibitor based on kinetic analysis.

Chinoketides A and B, Two New Antimicrobial Polyketides from the Endophytes of Distylium chinense with the “Black-Box” Co-culture Method

Two new polyketides, chinoketides A and B (1 – 2) with a known compound xylarphthalide A (3), were isolated from the solid medium of the endophytes from the leaves of the relic plant Distylium chinense with the “black-box” co-culture method, and the structures of two new compounds were elucidated by NMR, MS and CD spectra. And the absolute configurations of chinoketides A (1) and B (2) were determined as 2R,3R,8S and 5R,6S by calculating their ECD spectra to compare with the experimental CD spectra. Finally, the antimicrobial activities were evaluated to Erwinia carotovora sub sp. Carotovora (Jones) Bersey et al, and the results showed that compounds 1 – 3 displayed the antimicrobial activities with MIC value at 20.5, 30.4 and 10.2 µg/mL.

Screening of polyketide genes from Brazilian Atlantic Forest soil / Evaluación de genes biosintéticos de policétidos provenientes de suelo de la selva atlántica brasileñae / Avaliação de genes biossintéticos de policetídeos provenientes do solo da Mata Atlântica Brasileira

Abstract Soil is a large source of microorganisms with potential to produce bioactive compounds. Since most of them cannot be cultured, metagenomics has become a useful tool in order to evaluate this potential. The aim of this study was to screen biosynthetic polyketide genes (PKS) present in a metagenomic library constructed from a soil sample isolated from the Brazilian Atlantic Forest. The library comprises 5000 clones with DNA inserts between 40 and 50 Kb. The characterization of the biosynthetic gene clusters of these molecules is a promising alternative to elucidate the biotechnological potential of bioactive compounds in microbial communities. The PKS genes were screened using degenerated primers. The positive clones for PKS systems were isolated, and their nucleotide sequences analysed with bioinformatics tools. The screening yielded two positive clones for PKS II genes. Furthermore, variations in the sequences of the PKS II genes from the metagenomic library were observed when compared with sequences of ketosynthases' databases. With these findings we gain insight into the possible relation between new biosynthetic genes and the production of new secondary metabolites.

Resumen El suelo es una fuente importante de microrganismos con potencial para producir compuestos bioactivos. Dado que la gran mayoría de estos microorganismos no puede cultivarse, la metagenómica se ha convertido en una herramienta útil para evaluar dicho potencial. El objetivo del presente estudio fue evaluar los genes biosintéticos de policétidos (PKS) presentes en una biblioteca metagenómica construida a partir de una muestra de suelo aislada de la selva atlántica brasileña. La biblioteca comprende 5000 clones con insertos de DNA entre 40 y 50 Kb. La caracterización de clústeres de genes biosintéticos de estas moléculas es una alternativa promisoria para elucidar el potencial biotecnológico de los compuestos bioactivos en comunidades microbianas. Los genes biosintéticos de PKS se evaluaron usando cebadores degenerados. Se aislaron los clones positivos para sistemas PKS y sus secuencias de nucleótidos se analizaron con herramientas bioinformáticas. La evaluación arrojó dos clones positivos para genes de PKS II. Además, se observaron variaciones en las secuencias de genes de PKS II de la biblioteca metagenómica cuando se compararon con las secuencias de las bases de datos de cetosintasas. Estos hallazgos proporcionan nueva información sobre la posible relación entre nuevos genes biosintéticos y la producción de nuevos metabolitos secundarios.

Resumo O solo é uma fonte de importante de microrganismos com potencial para produzir compostos bioativos. Considerando que a maioria destes microrganismos não se pode cultivar, a metagenômica tem se convertido em uma ferramenta útil para avaliar este potencial. O objetivo deste estudo foi avaliar os genes biossintéticos de policetídeos (PKS) presentes em uma biblioteca metagenômica construída a partir de uma amostra de solo isolada da Mata Atlântica brasileira. A biblioteca compreende 5000 clones com insertos de DNA entre 40 e 50 Kb. A caracterização de clusters de genes biossintéticos destas moléculas é uma alternativa promissora para elucidar o potencial biotecnológico de compostos bioativos em comunidades microbianas. Os genes PKS foram avaliados usando primers degenerados. Os clones positivos para sistemas PKS foram isolados e suas sequências de nucleotídeos foram analisadas com ferramentas de bioinformática. A avaliação forneceu dois clones positivos para genes PKS II. Além disso, variações nas sequências dos genes PKS II da biblioteca metagenômica foram observadas quando comparadas com sequências da base de dados de cetosintases. Com estas descobertas obtivemos uma visão sobre uma possível relação entre novos genes biossintéticos e a produção de novos metabólitos secundários.

Bioprospección de la actividad antimicrobiana de los extractos de los hongos endófitos de Bauhinia guianensis / Bioprospecting of antimicrobial activity of extracts of endophytic fungi from Bauhinia guianensis

Antibiotic resistance results in higher medical costs, prolonged hospital stays and increased mortality and is rising to dangerously high levels in all parts of the world. Therefore, this study aims to search for new antimicrobial agents through bioprospecting of extracts of endophytic fungi from Bauhinia guianensis, a typical Amazonian plant used in combating infections. Seventeen (17) fungi were isolated and as result the methanolic extract of the fungus Exserohilum rostratum showed good activity against the bacteria tested. The polyketide monocerin was isolated by the chromatographic technique, identified by NMR and MS, showing broad antimicrobial spectrum.

La resistencia a los antibióticos conduce a mayores costos médicos, hospitalizaciones prolongadas e incremento de la mortalidad, y está aumentando a niveles peligrosamente altos en todas partes del mundo. Este estudio tuvo como objetivo la búsqueda de nuevos agentes antimicrobianos a través de la bioprospección de extractos de hongos endófitos de Bauhinia guianensis, una planta amazónica típica, utilizada en la lucha contra problemas infecciosos. Fueron aislados 17 hongos; el extracto metanólico del hongo Exserohilum rostratum mostró buena actividad contra las bacterias probadas. Se aisló monocerina policétido por la técnica de cromatografía; este compuesto fue identificado por RM y EM, y mostró un amplio espectro antimicrobiano.

Three New Non-reducing Polyketide Synthase Genes from the Lichen-Forming Fungus Usnea longissima

Usnea longissima has a long history of use as a traditional medicine. Several bioactive compounds, primarily belonging to the polyketide family, have been isolated from U. longissima. However, the genes for the biosynthesis of these compounds are yet to be identified. In the present study, three different types of non-reducing polyketide synthases (UlPKS2, UlPKS4, and UlPKS6) were identified from a cultured lichen-forming fungus of U. longissima. Phylogenetic analysis of product template domains showed that UlPKS2 and UlPKS4 belong to group IV, which includes the non-reducing polyketide synthases with an methyltransferase (MeT) domain that are involved in methylorcinol-based compound synthesis; UlPKS6 was found to belong to group I, which includes the non-reducing polyketide synthases that synthesize single aromatic ring polyketides, such as orsellinic acid. Reverse transcriptase-PCR analysis demonstrated that UlPKS2 and UlPKS4 were upregulated by sucrose; UlPKS6 was downregulated by asparagine, glycine, and alanine.

Biotechnology of polyketides: new breath of life for the novel antibiotic genetic pathways discovery through metagenomics

The discovery of secondary metabolites produced by microorganisms (e.g., penicillin in 1928) and the beginning of their industrial application (1940) opened new doors to what has been the main medication source for the treatment of infectious diseases and tumors. In fact, approximately 80 years after the discovery of the first antibiotic compound, and despite all of the warnings about the failure of the "goose that laid the golden egg," the potential of this wealth is still inexorable: simply adjust the focus from "micro" to "nano", that means changing the look from microorganisms to nanograms of DNA. Then, the search for new drugs, driven by genetic engineering combined with metagenomic strategies, shows us a way to bypass the barriers imposed by methodologies limited to isolation and culturing. However, we are far from solving the problem of supplying new molecules that are effective against the plasticity of multi- or pan-drug-resistant pathogens. Although the first advances in genetic engineering date back to 1990, there is still a lack of high-throughput methods to speed up the screening of new genes and design new molecules by recombination of pathways. In addition, it is necessary an increase in the variety of heterologous hosts and improvements throughout the full drug discovery pipeline. Among numerous studies focused on this subject, those on polyketide antibiotics stand out for the large technical-scientific efforts that established novel solutions for the transfer/engineering of major metabolic pathways using transposons and other episomes, overcoming one of the main methodological constraints for the heterologous expression of major pathways. In silico prediction analysis of three-dimensional enzymatic structures and advances in sequencing technologies have expanded access to the metabolic potential of microorganisms.

Computational Approach for the Analysis of Post-PKS Glycosylation Step

We introduce a computational approach for analysis of glycosylation in Post-PKS tailoring steps. It is a computational method to predict the deoxysugar biosynthesis unit pathway and the substrate specificity of glycosyltransferases involved in the glycosylation of polyketides. In this work, a directed and weighted graph is introduced to represent and predict the deoxysugar biosynthesis unit pathway. In addition, a homology based gene clustering method is used to predict the substrate specificity of glycosyltransferases. It is useful for the rational design of polyketide natural products, which leads to in silico drug discovery.