scholarly journals Diverse Protein Architectures and α-N-Methylation Patterns Define Split Borosin RiPP Biosynthetic Gene Clusters

2021 ◽  
Author(s):  
Aman S. Imani ◽  
Aileen R. Lee ◽  
Nisha Vishwanathan ◽  
Floris de Waal ◽  
Michael F. Freeman
Keyword(s):  
Markov Models ◽  
Sequence Similarity ◽  
Shewanella Oneidensis ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Peptide Backbone ◽  
Reading Frame ◽  
Type Iv ◽  
Modified Peptides

Borosins are ribosomally synthesized and post-translationally modified peptides (RiPPs) with α-N-methylations installed on the peptide backbone that impart unique properties like proteolytic stability to these natural products. The borosin RiPP family was initially reported only in fungi until our recent discovery and characterization of a Type IV split borosin system in the metal-respiring bacterium Shewanella oneidensis. Here, we used hidden Markov models and sequence similarity networks to identify over 1,600 putative pathways that show split borosin biosynthetic gene clusters are widespread in bacteria. Noteworthy differences in precursor and α-N-methyltransferase open reading frame sizes, architectures, and core peptide properties allow further subdivision of the borosin family into six additional discrete structural types, of which five have been validated in this study.

scholarly journals Pan-Genome of the Genus Streptomyces and Prioritization of Biosynthetic Gene Clusters With Potential to Produce Antibiotic Compounds

2021 ◽  
Vol 12 ◽  
Author(s):  
Carlos Caicedo-Montoya ◽  
Monserrat Manzo-Ruiz ◽  
Rigoberto Ríos-Estepa
Keyword(s):  
Comparative Genomics ◽  
Sequence Similarity ◽  
Experimental Studies ◽  
Gene Clusters ◽  
Antibiotic Activity ◽  
Genomic Diversity ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Genus Streptomyces ◽  
Pan Genome

Species of the genus Streptomyces are known for their ability to produce multiple secondary metabolites; their genomes have been extensively explored to discover new bioactive compounds. The richness of genomic data currently available allows filtering for high quality genomes, which in turn permits reliable comparative genomics studies and an improved prediction of biosynthetic gene clusters (BGCs) through genome mining approaches. In this work, we used 121 genome sequences of the genus Streptomyces in a comparative genomics study with the aim of estimating the genomic diversity by protein domains content, sequence similarity of proteins and conservation of Intergenic Regions (IGRs). We also searched for BGCs but prioritizing those with potential antibiotic activity. Our analysis revealed that the pan-genome of the genus Streptomyces is clearly open, with a high quantity of unique gene families across the different species and that the IGRs are rarely conserved. We also described the phylogenetic relationships of the analyzed genomes using multiple markers, obtaining a trustworthy tree whose relationships were further validated by Average Nucleotide Identity (ANI) calculations. Finally, 33 biosynthetic gene clusters were detected to have potential antibiotic activity and a predicted mode of action, which might serve up as a guide to formulation of related experimental studies.

scholarly journals Conformational rearrangements enable iterative backbone N-methylation in RiPP biosynthesis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Fredarla S. Miller ◽  
Kathryn K. Crone ◽  
Matthew R. Jensen ◽  
Sudipta Shaw ◽  
William R. Harcombe ◽  
...  
Keyword(s):  
Natural Product ◽  
Biological Membrane ◽  
Shewanella Oneidensis ◽  
Structural Constraints ◽  
Peptide Backbone ◽  
Amide Bonds ◽  
Type Iv ◽  
Precursor Peptide ◽  
Modified Peptides ◽  
Conformational Rearrangements

AbstractPeptide backbone α-N-methylations change the physicochemical properties of amide bonds to provide structural constraints and other favorable characteristics including biological membrane permeability to peptides. Borosin natural product pathways are the only known ribosomally encoded and posttranslationally modified peptides (RiPPs) pathways to incorporate backbone α-N-methylations on translated peptides. Here we report the discovery of type IV borosin natural product pathways (termed ‘split borosins’), featuring an iteratively acting α-N-methyltransferase and separate precursor peptide substrate from the metal-respiring bacterium Shewanella oneidensis. A series of enzyme-precursor complexes reveal multiple conformational states for both α-N-methyltransferase and substrate. Along with mutational and kinetic analyses, our results give rare context into potential strategies for iterative maturation of RiPPs.

scholarly journals Understanding Thioamitide Biosynthesis Using Pathway Engineering and Untargeted Metabolomics

2020 ◽  
Author(s):  
Tom H. Eyles ◽  
Natalia M. Vior ◽  
Rodney Lacret ◽  
Andrew W. Truman
Keyword(s):  
Biosynthetic Pathway ◽  
Gene Clusters ◽  
Untargeted Metabolomics ◽  
Pathway Engineering ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Heterologous Host ◽  
Detailed Understanding ◽  
Precursor Peptide ◽  
Modified Peptides

ABSTRACTThiostreptamide S4 is a thioamitide, a family of promising antitumour ribosomally synthesised and post-translationally modified peptides (RiPPs). The thioamitides are one of the most structurally complex RiPP families, yet very few thioamitide biosynthetic steps have been elucidated, even though the gene clusters of multiple thioamitides have been identified. We hypothesised that engineering the thiostreptamide S4 gene cluster in a heterologous host could provide insights into its biosynthesis when coupled with untargeted metabolomics and targeted mutations of the precursor peptide. Modified gene clusters were constructed, and in-depth metabolomics enabled a detailed understanding of the biosynthetic pathway, including the identification of an effector-like protein critical for amino acid dehydration. We use this biosynthetic understanding to bioinformatically identify new widespread families of RiPP biosynthetic gene clusters, paving the way for future RiPP discovery and engineering.

scholarly journals Streptomyces lydicamycinicus sp. nov. and Its Secondary Metabolite Biosynthetic Gene Clusters for Polyketide and Nonribosomal Peptide Compounds

2020 ◽  
Vol 8 (3) ◽  
pp. 370
Author(s):  
Hisayuki Komaki ◽  
Akira Hosoyama ◽  
Yasuhiro Igarashi ◽  
Tomohiko Tamura
Keyword(s):  
Sequence Similarity ◽  
Taxonomic Status ◽  
Gene Clusters ◽  
Diaminopimelic Acid ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Whole Genome Analysis ◽  
16S Rdna Sequence ◽  
Streptomyces Sp ◽  
Type Strains

(1) Background: Streptomyces sp. TP-A0598 derived from seawater produces lydicamycin and its congeners. We aimed to investigate its taxonomic status; (2) Methods: A polyphasic approach and whole genome analysis are employed; (3) Results: Strain TP-A0598 contained ll-diaminopimelic acid, glutamic acid, glycine, and alanine in its peptidoglycan. The predominant menaquinones were MK-9(H6) and MK-9(H8), and the major fatty acids were C16:0, iso-C15:0, iso-C16:0, and anteiso-C15:0. Streptomyces sp. TP-A0598 showed a 16S rDNA sequence similarity value of 99.93% (1 nucleottide difference) to Streptomyces angustmyceticus NRRL B-2347T. The digital DNA–DNA hybridisation value between Streptomyces sp. TP-A0598 and its closely related type strains was 25%–46%. Differences in phenotypic characteristics between Streptomyces sp. TP-A0598 and its phylogenetically closest relative, S. angustmyceticus NBRC 3934T, suggested strain TP-A0598 to be a novel species. Streptomyces sp. TP-A0598 and S. angustmyceticus NBRC 3934T harboured nine and 13 biosynthetic gene clusters for polyketides and nonribosomal peptides, respectively, among which only five clusters were shared between them, whereas the others are specific for each strain; and (4) Conclusions: For strain TP-A0598, the name Streptomyces lydicamycinicus sp. nov. is proposed; the type strain is TP-A0598T (=NBRC 110027T).

scholarly journals Recent development of antiSMASH and other computational approaches to mine secondary metabolite biosynthetic gene clusters

2017 ◽  
Vol 20 (4) ◽  
pp. 1103-1113 ◽  
Author(s):  
Kai Blin ◽  
Hyun Uk Kim ◽  
Marnix H Medema ◽  
Tilmann Weber
Keyword(s):  
Natural Products ◽  
Small Molecules ◽  
Sequence Similarity ◽  
Genome Mining ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Rule Based ◽  
Chemical Structures ◽  
Annotation Quality

Abstract Many drugs are derived from small molecules produced by microorganisms and plants, so-called natural products. Natural products have diverse chemical structures, but the biosynthetic pathways producing those compounds are often organized as biosynthetic gene clusters (BGCs) and follow a highly conserved biosynthetic logic. This allows for the identification of core biosynthetic enzymes using genome mining strategies that are based on the sequence similarity of the involved enzymes/genes. However, mining for a variety of BGCs quickly approaches a complexity level where manual analyses are no longer possible and require the use of automated genome mining pipelines, such as the antiSMASH software. In this review, we discuss the principles underlying the predictions of antiSMASH and other tools and provide practical advice for their application. Furthermore, we discuss important caveats such as rule-based BGC detection, sequence and annotation quality and cluster boundary prediction, which all have to be considered while planning for, performing and analyzing the results of genome mining studies.

scholarly journals Zn-dependent bifunctional proteases are responsible for leader peptide processing of class III lanthipeptides

2019 ◽  
Vol 116 (7) ◽  
pp. 2533-2538 ◽  
Author(s):  
Shaoming Chen ◽  
Bing Xu ◽  
Erquan Chen ◽  
Jiaqi Wang ◽  
Jingxia Lu ◽  
...  
Keyword(s):  
Gene Clusters ◽  
Leader Peptide ◽  
Biosynthetic Gene ◽  
Class Iii ◽  
Biosynthetic Gene Clusters ◽  
Peptide Processing ◽  
Leader Peptides ◽  
Biochemical Studies ◽  
Modified Peptides

Lanthipeptides are an important subfamily of ribosomally synthesized and posttranslationally modified peptides, and the removal of their N-terminal leader peptides by a designated protease(s) is a key step during maturation. Whereas proteases for class I and II lanthipeptides are well-characterized, the identity of the protease(s) responsible for class III leader processing remains unclear. Herein, we report that the class III lanthipeptide NAI-112 employs a bifunctional Zn-dependent protease, AplP, with both endo- and aminopeptidase activities to complete leader peptide removal, which is unprecedented in the biosynthesis of lanthipeptides. AplP displays a broad substrate scope in vitro by processing a number of class III leader peptides. Furthermore, our studies reveal that AplP-like proteases exist in the genomes of all class III lanthipeptide-producing strains but are usually located outside the biosynthetic gene clusters. Biochemical studies show that AplP-like proteases are universally responsible for the leader removal of the corresponding lanthipeptides. In addition, AplP-like proteases are phylogenetically correlated with aminopeptidase N from Escherichia coli, and might employ a single active site to catalyze both endo- and aminopeptidyl hydrolysis. These findings solve the long-standing question as to the mechanism of leader peptide processing during class III lanthipeptide biosynthesis, and pave the way for the production and bioengineering of this class of natural products.

scholarly journals Discovery and characterisation of an amidine-containing ribosomally-synthesised peptide that is widely distributed in nature

2021 ◽  
Author(s):  
Alicia H Russell ◽  
Natalia Miguel Vior ◽  
Edward Steven Hems ◽  
Rodney Lacret ◽  
Andrew William Truman
Keyword(s):  
Natural Product ◽  
Genome Mining ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Wide Range ◽  
Modified Peptides

Ribosomally synthesised and post-translationally modified peptides (RiPPs) are a structurally diverse class of natural product with a wide range of bioactivities. Genome mining for RiPP biosynthetic gene clusters (BGCs) is...

scholarly journals Discovery and characterisation of an amidine-containing ribosomally-synthesised peptide that is widely distributed in nature

2020 ◽  
Author(s):  
Alicia H. Russell ◽  
Natalia M. Vior ◽  
Edward S. Hems ◽  
Rodney Lacret ◽  
Andrew W. Truman
Keyword(s):  
Natural Product ◽  
Genome Mining ◽  
Gene Clusters ◽  
Model Organisms ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Post Translational Modifications ◽  
Streptomyces Albus ◽  
Modified Peptides ◽  
Mining Tool

ABSTRACTRibosomally synthesised and post-translationally modified peptides (RiPPs) are a structurally diverse class of natural product with a range of bioactivities. Genome mining for RiPP biosynthetic gene clusters (BGCs) is often hampered by poor detection of the short precursor peptides that are ultimately modified into the final molecule. Here, we utilise a previously described genome mining tool, RiPPER, to identify novel RiPP precursor peptides near YcaO-domain proteins, enzymes that catalyse various RiPP post-translational modifications including heterocyclisation and thioamidation. Using this dataset, we identified a novel, diverse and highly conserved family of RiPP BGCs spanning over 230 species of Actinobacteria and Firmicutes. A representative BGC from Streptomyces albus J1074 was characterised, leading to the discovery of streptamidine, a novel-amidine containing RiPP. This highlights the breadth of unexplored natural products with structurally rare features, even in model organisms.

scholarly journals Survey of Biosynthetic Gene Clusters from Sequenced Myxobacteria Reveals Unexplored Biosynthetic Potential

2019 ◽  
Vol 7 (6) ◽  
pp. 181 ◽  
Author(s):  
Katherine Gregory ◽  
Laura A. Salvador ◽  
Shukria Akbar ◽  
Barbara I. Adaikpoh ◽  
D. Cole Stevens
Keyword(s):  
Natural Product ◽  
Chemical Space ◽  
Sequence Similarity ◽  
Gene Clusters ◽  
Biologically Active ◽  
Critical Component ◽  
Biosynthetic Gene ◽  
Bacterial Genomes ◽  
Biosynthetic Gene Clusters ◽  
Natural Product Discovery

Coinciding with the increase in sequenced bacteria, mining of bacterial genomes for biosynthetic gene clusters (BGCs) has become a critical component of natural product discovery. The order Myxococcales, a reputable source of biologically active secondary metabolites, spans three suborders which all include natural product producing representatives. Utilizing the BiG-SCAPE-CORASON platform to generate a sequence similarity network that contains 994 BGCs from 36 sequenced myxobacteria deposited in the antiSMASH database, a total of 843 BGCs with lower than 75% similarity scores to characterized clusters within the MIBiG database are presented. This survey provides the biosynthetic diversity of these BGCs and an assessment of the predicted chemical space yet to be discovered. Considering the mere snapshot of myxobacteria included in this analysis, these untapped BGCs exemplify the potential for natural product discovery from myxobacteria.

scholarly journals Global analysis of adenylate-forming enzymes reveals β-lactone biosynthesis pathway in pathogenic Nocardia

2019 ◽  
Author(s):  
Serina L. Robinson ◽  
Barbara R. Terlouw ◽  
Megan D. Smith ◽  
Sacha J. Pidot ◽  
Tim P. Stinear ◽  
...  
Keyword(s):  
Machine Learning ◽  
Natural Product ◽  
Sequence Similarity ◽  
Global Analysis ◽  
Gene Clusters ◽  
Acid Activation ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Predictive Tool ◽  
Wide Range

ABSTRACTEnzymes that cleave ATP to activate carboxylic acids play essential roles in primary and secondary metabolism in all domains of life. Class I adenylate-forming enzymes share a conserved structural fold but act on a wide range of substrates to catalyze reactions involved in bioluminescence, nonribosomal peptide biosynthesis, fatty acid activation, and β-lactone formation. Despite their metabolic importance, the substrates and catalytic functions of the vast majority of adenylate-forming enzymes are unknown without tools available to accurately predict them. Given the crucial roles of adenylate-forming enzymes in biosynthesis, this also severely limits our ability to predict natural product structures from biosynthetic gene clusters. Here we used machine learning to predict adenylate-forming enzyme function and substrate specificity from protein sequence. We built a web-based predictive tool and used it to comprehensively map the biochemical diversity of adenylate-forming enzymes across >50,000 candidate biosynthetic gene clusters in bacterial, fungal, and plant genomes. Ancestral enzyme reconstruction and sequence similarity networking revealed a ‘hub’ topology suggesting radial divergence of the adenylate-forming superfamily from a core enzyme scaffold most related to contemporary aryl-CoA ligases. Our classifier also predicted β-lactone synthetases in novel biosynthetic gene clusters conserved across >90 different strains of Nocardia. To test our computational predictions, we purified a candidate β-lactone synthetase from Nocardia brasiliensis and reconstituted the biosynthetic pathway in vitro to link the gene cluster to the β-lactone natural product, nocardiolactone. We anticipate our machine learning approach will aid in functional classification of enzymes and advance natural product discovery.

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