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Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a large class of secondary metabolites that have garnered scientific attention due to their complex scaffolds with potential roles in medicine, agriculture, and chemical ecology. RiPPs derive from the cleavage of ribosomally synthesized proteins and additional modifications, catalyzed by various enzymes to alter the peptide backbone or side chains. Of these enzymes, cytochromes P450 (P450s) are a superfamily of heme-thiolate proteins involved in many metabolic pathways, including RiPP biosyntheses. In this review, we focus our discussion on P450 involved in RiPP pathways and the unique chemical transformations they mediate. Previous studies have revealed a wealth of P450s distributed across all domains of life. While the number of characterized P450s involved in RiPP biosyntheses is relatively small, they catalyze various enzymatic reactions such as C-C or C-N bond formation. Formation of some RiPPs is catalyzed by more than one P450, enabling structural diversity. With the continuous improvement of the bioinformatic tools for RiPP prediction and advancement in synthetic biology techniques, it is expected that further cytochrome P450-mediated RiPP biosynthetic pathways will be discovered.SummaryThe presence of genes encoding P450s in gene clusters for ribosomally synthesized and post-translationally modified peptides expand structural and functional diversity of these secondary metabolites, and here, we review the current state of this knowledge.

Streptomycetes are highly metabolically gifted bacteria with the abilities to produce bioproducts that have profound economic and societal importance. These bioproducts are produced by metabolic pathways including those for the biosynthesis of secondary metabolites and catabolism of plant biomass constituents. Advancements in genome sequencing technologies have revealed a wealth of untapped metabolic potential from Streptomyces genomes. Here, we report the largest Streptomyces pangenome generated by using 205 complete genomes. Metabolic potentials of the pangenome and individual genomes were analyzed, revealing degrees of conservation of individual metabolic pathways and strains potentially suitable for metabolic engineering. Of them, Streptomyces bingchenggensis was identified as a potent degrader of plant biomass. Polyketide, non-ribosomal peptide, and gamma-butyrolactone biosynthetic enzymes are primarily strain specific while ectoine and some terpene biosynthetic pathways are highly conserved. A large number of transcription factors associated with secondary metabolism are strain-specific while those controlling basic biological processes are highly conserved. Although the majority of genes involved in morphological development are highly conserved, there are strain-specific varieties which may contribute to fine tuning the timing of cellular differentiation. Overall, these results provide insights into the metabolic potential, regulation and physiology of streptomycetes, which will facilitate further exploitation of these important bacteria.

The vast majority of intracellular protein targets are refractory toward small-molecule therapeutic engagement, and additional therapeutic modalities are needed to overcome this deficiency. Here, the identification and characterization of a natural product, WDB002, reveals a therapeutic modality that dramatically expands the currently accepted limits of druggability. WDB002, in complex with the FK506-binding protein (FKBP12), potently and selectively binds the human centrosomal protein 250 (CEP250), resulting in disruption of CEP250 function in cells. The recognition mode is unprecedented in that the targeted domain of CEP250 is a coiled coil and is topologically featureless, embodying both a structural motif and surface topology previously considered on the extreme limits of "undruggability" for an intracellular target. Structural studies reveal extensive protein-WDB002 and protein-protein contacts, with the latter being distinct from those seen in FKBP12 ternary complexes formed by FK506 and rapamycin. Outward-facing structural changes in a bound small molecule can thus reprogram FKBP12 to engage diverse, otherwise "undruggable" targets. The flat-targeting modality demonstrated here has the potential to expand the druggable target range of small-molecule therapeutics. As CEP250 was recently found to be an interaction partner with the Nsp13 protein of the SARS-CoV-2 virus that causes COVID-19 disease, it is possible that WDB002 or an analog may exert useful antiviral activity through its ability to form high-affinity ternary complexes containing CEP250 and FKBP12.

AbstractMetabolomics is a widely used technology for obtaining direct measures of metabolic activities from diverse biological systems. However, ambiguous metabolite identifications are a common challenge and biochemical interpretation is often limited by incomplete and inaccurate genome-based predictions of enzyme activities (i.e. gene annotations). Metabolite, Annotation, and Gene Integration (MAGI) generates a metabolite-gene association score using a biochemical reaction network. This is calculated by a method that emphasizes consensus between metabolites and genes via biochemical reactions. To demonstrate the potential of this method, we applied MAGI to integrate sequence data and metabolomics data collected from Streptomyces coelicolor A3(2), an extensively characterized bacterium that produces diverse secondary metabolites. Our findings suggest that coupling metabolomics and genomics data by scoring consensus between the two increases the quality of both metabolite identifications and gene annotations in this organism. MAGI also made biochemical predictions for poorly annotated genes that were consistent with the extensive literature on this important organism. This limited analysis suggests that using metabolomics data has the potential to improve annotations in sequenced organisms and also provides testable hypotheses for specific biochemical functions. MAGI is freely available for academic use both as an online tool at https://magi.nersc.gov and with source code available at https://github.com/biorack/magi.

Genomic islands have been shown to harbor functional traits that differentiate ecologically distinct populations of environmental bacteria. A comparative analysis of the complete genome sequences of the marine Actinobacteria Salinispora tropica and Salinispora arenicola reveals that 75% of the species-specific genes are located in 21 genomic islands. These islands are enriched in genes associated with secondary metabolite biosynthesis providing evidence that secondary metabolism is linked to functional adaptation. Secondary metabolism accounts for 8.8% and 10.9% of the genes in the S. tropica and S. arenicola genomes, respectively, and represents the major functional category of annotated genes that differentiates the two species. Genomic islands harbor all 25 of the species-specific biosynthetic pathways, the majority of which occur in S. arenicola and may contribute to the cosmopolitan distribution of this species. Genome evolution is dominated by gene duplication and acquisition, which in the case of secondary metabolism provide immediate opportunities for the production of new bioactive products. Evidence that secondary metabolic pathways are exchanged horizontally, coupled with earlier evidence for fixation among globally distributed populations, supports a functional role and suggests that the acquisition of natural product biosynthetic gene clusters represents a previously unrecognized force driving bacterial diversification. Species-specific differences observed in clustered regularly interspaced short palindromic repeat sequences suggest that S. arenicola may possess a higher level of phage immunity, whereas a highly duplicated family of polymorphic membrane proteins provides evidence for a new mechanism of marine adaptation in Gram-positive bacteria.

Recent fermentation studies have identified actinomycetes of the marine-dwelling genus Salinispora as prolific natural product producers. To further evaluate their biosynthetic potential, we analyzed all identifiable secondary natural product gene clusters from the recently sequenced 5,184,724 bp S. tropica CNB-440 circular genome. Our analysis shows that biosynthetic potential meets or exceeds that shown by previous Streptomyces genome sequences as well as other natural product-producing actinomycetes. The S. tropica genome features nine polyketide synthase systems of every known formally classified family, non-ribosomal peptide synthetases and several hybrid clusters. While a few clusters appear to encode molecules previously identified in Streptomyces species, the majority of the 15 biosynthetic loci are novel. Specific chemical information about putative and observed natural product molecules is presented and discussed. In addition, our bioinformatic analysis was critical for the structure elucidation of the novel polyenemacrolactam salinilactam A. This study demonstrates the potential for genomic analysis to complement and strengthen traditional natural product isolation studies and firmly establishes the genus Salinispora as a rich source of novel drug-like molecules.

The isolation and structural characterization of two new diketopiperazine dipeptides, cyclomarazines A and B, and the new cyclic heptapeptide cyclomarin D from the marin bacterium Salinispora arenicola CNS-205 is described. The results provide insight into biosynthetic building blocks of cylomarin anti-inflammatory microbial peptides which is nonribosomally derived largely from nonproteinogenic amino acid residues.

Cytochrome p-450 mediated complex biosynthesis of aflatoxin B1 via the first oxidative cycle that result in the formation of 11-hydroxy o-methylsterigmatocystin (HOMST) is investigated by synthesizing HOMST by an alkylnitrilium variant of the Houben-Hoesch reaction. Results of the proposed hypothesis of HOMST formation and the mechanism of the second oxidative cycle of the P-450 are discussed.

A novel bacterial type III polyketide synthases (PKS) is identified that is required for germicidin biosynthesis, which appears to catalyze elongation of specific [beta]-ketoacyl-ACP thioester intermediates in fatty acid biosynthesis with ethyl- or methylmalonyl-CoA and subsequent cyclization of the resulting triketide. In vivo utilization of either ACP-tethered starter units or ethylmalonyl-CoA as an extender unit is unprecedented for any type III PKS, but only one type of III PKS is proposed for the biosynthesis of a natural product.

Recent advances in high-throughput DNA sequencing technologies have equipped biologists with a powerful new set of tools for advancing research goals. The resulting flood of sequence data has made it critically important to train the next generation of scientists to handle the inherent bioinformatic challenges. The North East Bioinformatics Collaborative (NEBC) is undertaking the genome sequencing and annotation of the little skate (Leucoraja erinacea) to promote advancement of bioinformatics infrastructure in our region, with an emphasis on practical education to create a critical mass of informatically savvy life scientists. In support of the Little Skate Genome Project, the NEBC members have developed several annotation workshops and jamborees to provide training in genome sequencing, annotation and analysis. Acting as a nexus for both curation activities and dissemination of project data, a project web portal, SkateBase (http://skatebase.org) has been developed. As a case study to illustrate effective coupling of community annotation with workforce development, we report the results of the Mitochondrial Genome Annotation Jamborees organized to annotate the first completely assembled element of the Little Skate Genome Project, as a culminating experience for participants from our three prior annotation workshops. We are applying the physical/virtual infrastructure and lessons learned from these activities to enhance and streamline the genome annotation workflow, as we look toward our continuing efforts for larger-scale functional and structural community annotation of the L. erinacea genome. [ABSTRACT FROM AUTHOR]

The article cites a study which identified a novel bacterial type III polyketide synthases (PKS). The novel bacteria is required for germicidin assembly that appears to catalyze elongation of specific β-ketoacyl-ACP thioester intermediates in fatty acid biosynthesis with ethyl- and methylmalonyl-CoA enzyme. Researchers used a Streptomyces coelicolor-based genome mining to identify this bacteria.

Polyketide synthases (PKSs) are involved in the biosynthesis of many important natural products. In bacteria, type III PKSs typically catalyze iterative decarboxylation and condensation reactions of malonyl-CoA building blocks in the biosynthesis of polyhydroxyaromatic products. Here it is shown that Gcs, a type III PKS encoded by the sco7221 ORF of the bacterium Streptomyces coelicolor, is required for biosynthesis of the germicidin family of 3,6-dialkyl-4-hydroxypyran-2-one natural products. Evidence consistent with Gcs-catalyzed elongation of specific beta-ketoacyl-ACP products of the fatty acid synthase FabH with ethyl- or methylmalonyl-CoA in the biosynthesis of germicidins is presented. Selectivity for beta-ketoacyl-ACP starter units and ethylmalonyl-CoA as an extender unit is unprecedented for type III PKSs, suggesting these enzymes may be capable of utilizing a far wider range of starter and extender units for natural product assembly than believed until now.