Genomic and chemical decryption of the Bacteroidetes phylum for its potential to biosynthesize natural products

With progress in genome sequencing and data sharing, 1000s of bacterial genomes are publicly available. Genome mining – using bioinformatics tools in terms of biosynthetic gene cluster (BGC) identification, analysis and rating – has become a key technology to explore the capabilities for natural product (NP) biosynthesis. Comprehensively, analyzing the genetic potential of the phylum Bacteroidetes revealed Chitinophaga as the most talented genus in terms of BGC abundance and diversity. Guided by the computational predictions, we conducted a metabolomics and bioactivity driven NP discovery program on 25 Chitinophaga strains. High numbers of peerless strain-specific metabolite buckets confirmed the upfront predicted biosynthetic potential and revealed a tremendous uncharted chemical space. Sourcing this dataset, we isolated the new iron chelating nonribosomally synthesized cyclic tetradeca- and pentadecalipodepsipeptide antibiotics chitinopeptins with activity against Candida, produced by C. eiseniae DSM 22224 and C. flava KCTC 62435, respectively.

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Recapitulation of the evolution of biosynthetic gene clusters reveals hidden chemical diversity on bacterial genomes

10.1101/020503 ◽

2015 ◽

Cited By ~ 6

Author(s):

Pablo Cruz-Morales ◽

Christian E. Martínez-Guerrero ◽

Marco A. Morales-Escalante ◽

Luis Yáñez-Guerra ◽

Johannes Florian Kopp ◽

...

Keyword(s):

Natural Products ◽

Chemical Space ◽

Streptomyces Coelicolor ◽

Genome Mining ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Chemical Diversity ◽

Biosynthetic Gene ◽

Bacterial Genomes ◽

Biosynthetic Gene Clusters

AbstractNatural products have provided humans with antibiotics for millennia. However, a decline in the pace of chemical discovery exerts pressure on human health as antibiotic resistance spreads. The empirical nature of current genome mining approaches used for natural products research limits the chemical space that is explored. By integration of evolutionary concepts related to emergence of metabolism, we have gained fundamental insights that are translated into an alternative genome mining approach, termed EvoMining. As the founding assumption of EvoMining is the evolution of enzymes, we solved two milestone problems revealing unprecedented conversions. First, we report the biosynthetic gene cluster of the ‘orphan’ metabolite leupeptin in Streptomyces roseus. Second, we discover an enzyme involved in formation of an arsenic-carbon bond in Streptomyces coelicolor and Streptomyces lividans. This work provides evidence that bacterial chemical repertoire is underexploited, as well as an approach to accelerate the discovery of novel antibiotics from bacterial genomes.

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Strain Prioritization and Genome Mining for Enediyne Natural Products

mBio ◽

10.1128/mbio.02104-16 ◽

2016 ◽

Vol 7 (6) ◽

Cited By ~ 45

Author(s):

Xiaohui Yan ◽

Huiming Ge ◽

Tingting Huang ◽

Hindra ◽

Dong Yang ◽

...

Keyword(s):

Natural Products ◽

Cancer Cell ◽

Polyketide Synthase ◽

Chemical Space ◽

Large Strain ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Structural Features ◽

A Genome ◽

Strain Collection

ABSTRACT The enediyne family of natural products has had a profound impact on modern chemistry, biology, and medicine, and yet only 11 enediynes have been structurally characterized to date. Here we report a genome survey of 3,400 actinomycetes, identifying 81 strains that harbor genes encoding the enediyne polyketide synthase cassettes that could be grouped into 28 distinct clades based on phylogenetic analysis. Genome sequencing of 31 representative strains confirmed that each clade harbors a distinct enediyne biosynthetic gene cluster. A genome neighborhood network allows prediction of new structural features and biosynthetic insights that could be exploited for enediyne discovery. We confirmed one clade as new C-1027 producers, with a significantly higher C-1027 titer than the original producer, and discovered a new family of enediyne natural products, the tiancimycins (TNMs), that exhibit potent cytotoxicity against a broad spectrum of cancer cell lines. Our results demonstrate the feasibility of rapid discovery of new enediynes from a large strain collection. IMPORTANCE Recent advances in microbial genomics clearly revealed that the biosynthetic potential of soil actinomycetes to produce enediynes is underappreciated. A great challenge is to develop innovative methods to discover new enediynes and produce them in sufficient quantities for chemical, biological, and clinical investigations. This work demonstrated the feasibility of rapid discovery of new enediynes from a large strain collection. The new C-1027 producers, with a significantly higher C-1027 titer than the original producer, will impact the practical supply of this important drug lead. The TNMs, with their extremely potent cytotoxicity against various cancer cells and their rapid and complete cancer cell killing characteristics, in comparison with the payloads used in FDA-approved antibody-drug conjugates (ADCs), are poised to be exploited as payload candidates for the next generation of anticancer ADCs. Follow-up studies on the other identified hits promise the discovery of new enediynes, radically expanding the chemical space for the enediyne family.

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Biosynthetic potential of uncultured Antarctic soil bacteria revealed through long-read metagenomic sequencing

The ISME Journal ◽

10.1038/s41396-021-01052-3 ◽

2021 ◽

Author(s):

Valentin Waschulin ◽

Chiara Borsetto ◽

Robert James ◽

Kevin K. Newsham ◽

Stefano Donadio ◽

...

Keyword(s):

Genome Mining ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Full Length ◽

Metagenomic Sequencing ◽

Short Read ◽

Short Read Sequencing ◽

Rich Diversity ◽

Long Read ◽

The Rich

AbstractThe growing problem of antibiotic resistance has led to the exploration of uncultured bacteria as potential sources of new antimicrobials. PCR amplicon analyses and short-read sequencing studies of samples from different environments have reported evidence of high biosynthetic gene cluster (BGC) diversity in metagenomes, indicating their potential for producing novel and useful compounds. However, recovering full-length BGC sequences from uncultivated bacteria remains a challenge due to the technological restraints of short-read sequencing, thus making assessment of BGC diversity difficult. Here, long-read sequencing and genome mining were used to recover >1400 mostly full-length BGCs that demonstrate the rich diversity of BGCs from uncultivated lineages present in soil from Mars Oasis, Antarctica. A large number of highly divergent BGCs were not only found in the phyla Acidobacteriota, Verrucomicrobiota and Gemmatimonadota but also in the actinobacterial classes Acidimicrobiia and Thermoleophilia and the gammaproteobacterial order UBA7966. The latter furthermore contained a potential novel family of RiPPs. Our findings underline the biosynthetic potential of underexplored phyla as well as unexplored lineages within seemingly well-studied producer phyla. They also showcase long-read metagenomic sequencing as a promising way to access the untapped genetic reservoir of specialised metabolite gene clusters of the uncultured majority of microbes.

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Discovery of a Potential Liver Fibrosis Inhibitor from a Mushroom Endophytic Fungus by Genome Mining of a Silent Biosynthetic Gene Cluster

Journal of Agricultural and Food Chemistry ◽

10.1021/acs.jafc.1c03639 ◽

2021 ◽

Vol 69 (38) ◽

pp. 11303-11310

Author(s):

Jin-Tao Cheng ◽

Han-Min Wang ◽

Jia-Hui Yu ◽

Chen-Fan Sun ◽

Fei Cao ◽

...

Keyword(s):

Liver Fibrosis ◽

Gene Cluster ◽

Endophytic Fungus ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene

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Analysis of the complete genome sequence of a marine-derived strainStreptomycessp. S063 CGMCC 14582 reveals its biosynthetic potential to produce novel anti-complement agents and peptides

PeerJ ◽

10.7717/peerj.6122 ◽

2019 ◽

Vol 7 ◽

pp. e6122 ◽

Cited By ~ 2

Author(s):

Liang-Yu Chen ◽

Hao-Tian Cui ◽

Chun Su ◽

Feng-Wu Bai ◽

Xin-Qing Zhao

Keyword(s):

Bioactive Compounds ◽

Genome Sequence ◽

Complete Genome Sequence ◽

Dna Hybridization ◽

Complete Genome ◽

System Analysis ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Genome Comparison ◽

Genome Sequences

Genome sequences of marine streptomycetes are valuable for the discovery of useful enzymes and bioactive compounds by genome mining. However, publicly available complete genome sequences of marine streptomycetes are still limited. Here, we present the complete genome sequence of a marine streptomyceteStreptomycessp. S063 CGMCC 14582. Species delineation based on the pairwise digital DNA-DNA hybridization and genome comparison ANI (average nucleotide identity) value showed thatStreptomycessp. S063 CGMCC 14582 possesses a unique genome that is clearly different from all of the other available genomes. Bioactivity tests showed thatStreptomycessp. S063 CGMCC 14582 produces metabolites with anti-complement activities, which are useful for treatment of numerous diseases that arise from inappropriate activation of the human complement system. Analysis of the genome reveals no biosynthetic gene cluster (BGC) which shows even low similarity to that of the known anti-complement agents was detected in the genome, indicating thatStreptomycessp. S063 CGMCC 14582 may produce novel anti-complement agents of microbial origin. Four BGCs which are potentially involved in biosynthesis of non-ribosomal peptides were disrupted, but no decrease of anti-complement activities was observed, suggesting that these four BGCs are not involved in biosynthesis of the anti-complement agents. In addition, LC-MS/MS analysis and subsequent alignment through the Global Natural Products Social Molecular Networking (GNPS) platform led to the detection of novel peptides produced by the strain.Streptomycessp. S063 CGMCC 14582 grows rapidly and is salt tolerant, which benefits efficient secondary metabolite production via seawater-based fermentation. Our results indicate thatStreptomycessp. S063 has great potential to produce novel bioactive compounds, and also is a good host for heterologous production of useful secondary metabolites for drug discovery.

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Discovery of Unusual Cyanobacterial Tryptophan-containing Anabaenopeptins by MS/MS Based Molecular Networking

10.20944/preprints202007.0562.v1 ◽

2020 ◽

Author(s):

Subhasish Saha ◽

Germana Esposito ◽

Petra Urajova ◽

Jan Mareš ◽

Daniela Ewe ◽

...

Keyword(s):

Multidisciplinary Approach ◽

Spectroscopic Analysis ◽

Chemical Space ◽

Genome Mining ◽

Gc Content ◽

Gene Clusters ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Hela Cell Lines ◽

Bioactive Secondary Metabolites

Heterocytous cyanobacteria are among the most prolific source of bioactive secondary metabolites, including anabaenopeptins (APTs). A terrestrial filamentous Brasilonema sp. CT11 collected in Costa Rica bamboo forest, as black mat was studied using a multidisciplinary approach: genome mining and HPLC-HRMS/MS coupled with bionformatic analyses. Herein, we report the nearly complete genome consisting 8.79 Mbp with a GC content of 42.4%. Moreover, we report on three novel tryptophane-containing APTs; anabaenopeptin 788 (1), anabaenopeptin 802 (2) and anabaenopeptin 816 (3). Further, the structure of two homologues, i.e., anabaenopeptin 802 (2a) and anabaenopeptin 802 (2b) was determined by spectroscopic analysis (NMR and MS). Both compounds were shown to exert weak to moderate antiproliferative activity against HeLa cell lines. This study also provides the unique and diverse potential of biosynthetic gene clusters and an assessment of the predicted chemical space yet to be discovered from this genus.

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Genome Mining Coupled with OSMAC-Based Cultivation Reveal Differential Production of Surugamide A by the Marine Sponge Isolate Streptomyces sp. SM17 When Compared to Its Terrestrial Relative S. albidoflavus J1074

Microorganisms ◽

10.3390/microorganisms7100394 ◽

2019 ◽

Vol 7 (10) ◽

pp. 394 ◽

Cited By ~ 5

Author(s):

Eduardo Almeida ◽

Navdeep Kaur ◽

Laurence Jennings ◽

Andrés Felipe Carrillo Rincón ◽

Stephen Jackson ◽

...

Keyword(s):

Marine Sponge ◽

Carbon Catabolite Repression ◽

Marine Invertebrates ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene ◽

Streptomyces Sp ◽

Streptomyces Albidoflavus ◽

Recent Interest ◽

New Compounds

Much recent interest has arisen in investigating Streptomyces isolates derived from the marine environment in the search for new bioactive compounds, particularly those found in association with marine invertebrates, such as sponges. Among these new compounds recently identified from marine Streptomyces isolates are the octapeptidic surugamides, which have been shown to possess anticancer and antifungal activities. By employing genome mining followed by an one strain many compounds (OSMAC)-based approach, we have identified the previously unreported capability of a marine sponge-derived isolate, namely Streptomyces sp. SM17, to produce surugamide A. Phylogenomics analyses provided novel insights on the distribution and conservation of the surugamides biosynthetic gene cluster (sur BGC) and suggested a closer relatedness between marine-derived sur BGCs than their terrestrially derived counterparts. Subsequent analysis showed differential production of surugamide A when comparing the closely related marine and terrestrial isolates, namely Streptomyces sp. SM17 and Streptomyces albidoflavus J1074. SM17 produced higher levels of surugamide A than S. albidoflavus J1074 under all conditions tested, and in particular producing >13-fold higher levels when grown in YD and 3-fold higher levels in SYP-NaCl medium. In addition, surugamide A production was repressed in TSB and YD medium, suggesting that carbon catabolite repression (CCR) may influence the production of surugamides in these strains.

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Pyridoxal-5′-phosphate–dependent bifunctional enzyme catalyzed biosynthesis of indolizidine alkaloids in fungi

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1914777117 ◽

2019 ◽

Vol 117 (2) ◽

pp. 1174-1180 ◽

Cited By ~ 3

Author(s):

Guang Zhi Dai ◽

Wen Bo Han ◽

Ya Ning Mei ◽

Kuang Xu ◽

Rui Hua Jiao ◽

...

Keyword(s):

Polyketide Synthase ◽

Complex Structure ◽

Genome Mining ◽

Biosynthetic Gene Cluster ◽

Major Facilitator Superfamily ◽

Molecular Architecture ◽

Widespread Occurrence ◽

Indolizidine Alkaloids ◽

Major Facilitator ◽

Insight Into

Indolizidine alkaloids such as anticancer drugs vinblastine and vincristine are exceptionally attractive due to their widespread occurrence, prominent bioactivity, complex structure, and sophisticated involvement in the chemical defense for the producing organisms. However, the versatility of the indolizidine alkaloid biosynthesis remains incompletely addressed since the knowledge about such biosynthetic machineries is only limited to several representatives. Herein, we describe the biosynthetic gene cluster (BGC) for the biosynthesis of curvulamine, a skeletally unprecedented antibacterial indolizidine alkaloid from Curvularia sp. IFB-Z10. The molecular architecture of curvulamine results from the functional collaboration of a highly reducing polyketide synthase (CuaA), a pyridoxal-5′-phosphate (PLP)-dependent aminotransferase (CuaB), an NADPH-dependent dehydrogenase (CuaC), and a FAD-dependent monooxygenase (CuaD), with its transportation and abundance regulated by a major facilitator superfamily permease (CuaE) and a Zn(II)Cys6 transcription factor (CuaF), respectively. In contrast to expectations, CuaB is bifunctional and capable of catalyzing the Claisen condensation to form a new C–C bond and the α-hydroxylation of the alanine moiety in exposure to dioxygen. Inspired and guided by the distinct function of CuaB, our genome mining effort discovers bipolamines A−I (bipolamine G is more antibacterial than curvulamine), which represent a collection of previously undescribed polyketide alkaloids from a silent BGC in Bipolaris maydis ATCC48331. The work provides insight into nature’s arsenal for the indolizidine-coined skeletal formation and adds evidence in support of the functional versatility of PLP-dependent enzymes in fungi.

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Genomic analysis of siderophore β-hydroxylases reveals divergent stereocontrol and expands the condensation domain family

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1903161116 ◽

2019 ◽

Vol 116 (40) ◽

pp. 19805-19814 ◽

Cited By ~ 8

Author(s):

Zachary L. Reitz ◽

Clifford D. Hardy ◽

Jaewon Suk ◽

Jean Bouvet ◽

Alison Butler

Keyword(s):

Predictive Power ◽

Genome Mining ◽

Genomic Analysis ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Peptide Synthetase ◽

Condensation Domain

Genome mining of biosynthetic pathways streamlines discovery of secondary metabolites but can leave ambiguities in the predicted structures, which must be rectified experimentally. Through coupling the reactivity predicted by biosynthetic gene clusters with verified structures, the origin of the β-hydroxyaspartic acid diastereomers in siderophores is reported herein. Two functional subtypes of nonheme Fe(II)/α-ketoglutarate–dependent aspartyl β-hydroxylases are identified in siderophore biosynthetic gene clusters, which differ in genomic organization—existing either as fused domains (IβHAsp) at the carboxyl terminus of a nonribosomal peptide synthetase (NRPS) or as stand-alone enzymes (TβHAsp)—and each directs opposite stereoselectivity of Asp β-hydroxylation. The predictive power of this subtype delineation is confirmed by the stereochemical characterization of β-OHAsp residues in pyoverdine GB-1, delftibactin, histicorrugatin, and cupriachelin. The l-threo (2S, 3S) β-OHAsp residues of alterobactin arise from hydroxylation by the β-hydroxylase domain integrated into NRPS AltH, while l-erythro (2S, 3R) β-OHAsp in delftibactin arises from the stand-alone β-hydroxylase DelD. Cupriachelin contains both l-threo and l-erythro β-OHAsp, consistent with the presence of both types of β-hydroxylases in the biosynthetic gene cluster. A third subtype of nonheme Fe(II)/α-ketoglutarate–dependent enzymes (IβHHis) hydroxylates histidyl residues with l-threo stereospecificity. A previously undescribed, noncanonical member of the NRPS condensation domain superfamily is identified, named the interface domain, which is proposed to position the β-hydroxylase and the NRPS-bound amino acid prior to hydroxylation. Through mapping characterized β-OHAsp diastereomers to the phylogenetic tree of siderophore β-hydroxylases, methods to predict β-OHAsp stereochemistry in silico are realized.

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