In Silico Analysis of PKS and NRPS Gene Clusters in Arisostatin- and Kosinostatin-Producers and Description of Micromonospora okii sp. nov.

Micromonospora sp. TP-A0316 and Micromonospora sp. TP-A0468 are producers of arisostatin and kosinostatin, respectively. Micromonospora sp. TP-A0316 showed a 16S rRNA gene sequence similarity of 100% to Micromonosporaoryzae CP2R9-1T whereas Micromonospora sp. TP-A0468 showed a 99.3% similarity to Micromonospora haikouensis 232617T. A phylogenetic analysis based on gyrB sequences suggested that Micromonospora sp. TP-A0316 is closely related to Micromonospora oryzae whereas Micromonospora TP-A0468 is an independent genomospecies. As Micromonospora sp. TP-A0468 showed some phenotypic differences to its closely related species, it was classified as a novel species, for which the name Micromonospora okii sp. nov. is proposed. The type strain is TP-A0468T (= NBRC 110461T). Micromonospora sp. TP-A0316 and M. okii TP-A0468T were both found to harbor 15 gene clusters for secondary metabolites such as polyketides and nonribosomal peptides in their genomes. Arisostatin-biosynthetic gene cluster (BGC) of Micromonospora sp. TP-A0316 closely resembled tetrocarcin A-BGC of Micromonospora chalcea NRRL 11289. A large type-I polyketide synthase gene cluster was present in each genome of Micromonospora sp. TP-A0316 and M. okii TP-A0468T. It was an ortholog of quinolidomicin-BGC of M. chalcea AK-AN57 and widely distributed in the genus Micromonospora.

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Microfluidic automated plasmid library enrichment for biosynthetic gene cluster discovery

Nucleic Acids Research ◽

10.1093/nar/gkaa131 ◽

2020 ◽

Vol 48 (8) ◽

pp. e48-e48 ◽

Cited By ~ 1

Author(s):

Peng Xu ◽

Cyrus Modavi ◽

Benjamin Demaree ◽

Frederick Twigg ◽

Benjamin Liang ◽

...

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Bioactive Molecules ◽

Type I ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Plasmid Library ◽

Polyketide Synthase Gene

Abstract Microbial biosynthetic gene clusters are a valuable source of bioactive molecules. However, because they typically represent a small fraction of genomic material in most metagenomic samples, it remains challenging to deeply sequence them. We present an approach to isolate and sequence gene clusters in metagenomic samples using microfluidic automated plasmid library enrichment. Our approach provides deep coverage of the target gene cluster, facilitating reassembly. We demonstrate the approach by isolating and sequencing type I polyketide synthase gene clusters from an Antarctic soil metagenome. Our method promotes the discovery of functional-related genes and biosynthetic pathways.

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The Fusarium verticillioides FUM Gene Cluster Encodes a Zn(II)2Cys6 Protein That Affects FUM Gene Expression and Fumonisin Production

Eukaryotic Cell ◽

10.1128/ec.00400-06 ◽

2007 ◽

Vol 6 (7) ◽

pp. 1210-1218 ◽

Cited By ~ 137

Author(s):

Daren W. Brown ◽

Robert A. E. Butchko ◽

Mark Busman ◽

Robert H. Proctor

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Fusarium Verticillioides ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Regulatory Proteins ◽

Wild Type ◽

Polyketide Synthase Gene ◽

Splice Forms ◽

Self Protection

ABSTRACT Fumonisins are mycotoxins produced by some Fusarium species and can contaminate maize or maize products. Ingestion of fumonisins is associated with diseases, including cancer and neural tube defects, in humans and animals. In fungi, genes involved in the synthesis of mycotoxins and other secondary metabolites are often located adjacent to each other in gene clusters. Such genes can encode structural enzymes, regulatory proteins, and/or proteins that provide self-protection. The fumonisin biosynthetic gene cluster includes 16 genes, none of which appear to play a role in regulation. In this study, we identified a previously undescribed gene (FUM21) located adjacent to the fumonisin polyketide synthase gene, FUM1. The presence of a Zn(II)2Cys6 DNA-binding domain in the predicted protein suggested that FUM21 was involved in transcriptional regulation. FUM21 deletion (Δfum21) mutants produce little to no fumonisin in cracked maize cultures but some FUM1 and FUM8 transcripts in a liquid GYAM medium. Complementation of a Δfum21 mutant with a wild-type copy of the gene restored fumonisin production. Analysis of FUM21 cDNAs identified four alternative splice forms (ASFs), and microarray analysis indicated the ASFs were differentially expressed. Based on these data, we present a model for how FUM21 ASFs may regulate fumonisin biosynthesis.

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Genome Sequence-Guided Finding of Lucensomycin Production by Streptomyces achromogenes Subsp. streptozoticus NBRC14001

Microorganisms ◽

10.3390/microorganisms10010037 ◽

2021 ◽

Vol 10 (1) ◽

pp. 37

Author(s):

Sho Nishimura ◽

Kazune Nakamura ◽

Miyako Yamamoto ◽

Daichi Morita ◽

Teruo Kuroda ◽

...

Keyword(s):

Antifungal Activity ◽

Gene Cluster ◽

Genome Sequence ◽

Polyketide Synthase ◽

Macrolide Antibiotic ◽

Biosynthetic Gene Cluster ◽

Antifungal Compound ◽

Type I ◽

Biosynthetic Gene ◽

Polyketide Synthase Gene

Information on microbial genome sequences is a powerful resource for accessing natural products with significant activities. We herein report the unveiling of lucensomycin production by Streptomyces achromogenes subsp. streptozoticus NBRC14001 based on the genome sequence of the strain. The genome sequence of strain NBRC14001 revealed the presence of a type I polyketide synthase gene cluster with similarities to a biosynthetic gene cluster for natamycin, which is a polyene macrolide antibiotic that exhibits antifungal activity. Therefore, we investigated whether strain NBRC14001 produces antifungal compound(s) and revealed that an extract from the strain inhibited the growth of Candida albicans. A HPLC analysis of a purified compound exhibiting antifungal activity against C. albicans showed that the compound differed from natamycin. Based on HR-ESI-MS spectrometry and a PubChem database search, the compound was predicted to be lucensomycin, which is a tetraene macrolide antibiotic, and this prediction was supported by the results of a MS/MS analysis. Furthermore, the type I polyketide synthase gene cluster in strain NBRC14001 corresponded well to lucesomycin biosynthetic gene cluster (lcm) in S. cyanogenus, which was very recently reported. Therefore, we concluded that the antifungal compound produced by strain NBRC14001 is lucensomycin.

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Characterization of the Nodularin Synthetase Gene Cluster and Proposed Theory of the Evolution of Cyanobacterial Hepatotoxins

Applied and Environmental Microbiology ◽

10.1128/aem.70.11.6353-6362.2004 ◽

2004 ◽

Vol 70 (11) ◽

pp. 6353-6362 ◽

Cited By ~ 169

Author(s):

Michelle C. Moffitt ◽

Brett A. Neilan

Keyword(s):

Gene Cluster ◽

Rational Design ◽

Polyketide Synthase ◽

Alternative Hypothesis ◽

Phylogenetic Analyses ◽

Cyanobacterial Bloom ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Biosynthetic Gene ◽

Microcystin Synthetase

ABSTRACT Nodularia spumigena is a bloom-forming cyanobacterium which produces the hepatotoxin nodularin. The complete gene cluster encoding the enzymatic machinery required for the biosynthesis of nodularin in N. spumigena strain NSOR10 was sequenced and characterized. The 48-kb gene cluster consists of nine open reading frames (ORFs), ndaA to ndaI, which are transcribed from a bidirectional regulatory promoter region and encode nonribosomal peptide synthetase modules, polyketide synthase modules, and tailoring enzymes. The ORFs flanking the nda gene cluster in the genome of N. spumigena strain NSOR10 were identified, and one of them was found to encode a protein with homology to previously characterized transposases. Putative transposases are also associated with the structurally related microcystin synthetase (mcy) gene clusters derived from three cyanobacterial strains, indicating a possible mechanism for the distribution of these biosynthetic gene clusters between various cyanobacterial genera. We propose an alternative hypothesis for hepatotoxin evolution in cyanobacteria based on the results of comparative and phylogenetic analyses of the nda and mcy gene clusters. These analyses suggested that nodularin synthetase evolved from a microcystin synthetase progenitor. The identification of the nodularin biosynthetic gene cluster and evolution of hepatotoxicity in cyanobacteria reported in this study may be valuable for future studies on toxic cyanobacterial bloom formation. In addition, an appreciation of the natural evolution of nonribosomal biosynthetic pathways will be vital for future combinatorial engineering and rational design of novel metabolites and pharmaceuticals.

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Characterization of the Azinomycin B Biosynthetic Gene Cluster Revealing a Different Iterative Type I Polyketide Synthase for Naphthoate Biosynthesis

Chemistry & Biology ◽

10.1016/j.chembiol.2008.05.021 ◽

2008 ◽

Vol 15 (7) ◽

pp. 693-705 ◽

Cited By ~ 74

Author(s):

Qunfei Zhao ◽

Qingli He ◽

Wei Ding ◽

Mancheng Tang ◽

Qianjin Kang ◽

...

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Biosynthetic Gene Cluster ◽

Type I ◽

Biosynthetic Gene ◽

Azinomycin B

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Cloning, Sequencing, and Functional Analysis of an Iterative Type I Polyketide Synthase Gene Cluster for Biosynthesis of the Antitumor Chlorinated Polyenone Neocarzilin in “Streptomyces carzinostaticus”

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.9.3468-3476.2004 ◽

2004 ◽

Vol 48 (9) ◽

pp. 3468-3476 ◽

Cited By ~ 43

Author(s):

Miyuki Otsuka ◽

Koji Ichinose ◽

Isao Fujii ◽

Yutaka Ebizuka

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Streptomyces Coelicolor ◽

Open Reading Frames ◽

Modular Organization ◽

Type I ◽

Orf3 Protein ◽

Polyketide Synthase Gene ◽

Polyketide Chain ◽

Reading Frames

ABSTRACT Neocarzilins (NCZs) are antitumor chlorinated polyenones produced by “Streptomyces carzinostaticus” var. F-41. The gene cluster responsible for the biosynthesis of NCZs was cloned and characterized. DNA sequence analysis of a 33-kb region revealed a cluster of 14 open reading frames (ORFs), three of which (ORF4, ORF5, and ORF6) encode type I polyketide synthase (PKS), which consists of four modules. Unusual features of the modular organization is the lack of an obvious acyltransferase domain on modules 2 and 4 and the presence of longer interdomain regions more than 200 amino acids in length on each module. Involvement of the PKS genes in NCZ biosynthesis was demonstrated by heterologous expression of the cluster in Streptomyces coelicolor CH999, which produced the apparent NCZ biosynthetic intermediates dechloroneocarzillin A and dechloroneocarzilin B. Disruption of ORF5 resulted in a failure of NCZ production, providing further evidence that the cluster is essential for NCZ biosynthesis. Mechanistic consideration of NCZ formation indicates the iterative use of at least one module of the PKS, which subsequently releases its product by decarboxylation to generate an NCZ skeleton, possibly catalyzed by a type II thioesterase encoded by ORF7. This is a novel type I PKS system of bacterial origin for the biosynthesis of a reduced polyketide chain. Additionally, the protein encoded by ORF3, located upstream of the PKS genes, closely resembles the FADH2-dependent halogenases involved in the formation of halometabolites. The ORF3 protein could be responsible for the halogenation of NCZs, presenting a unique example of a halogenase involved in the biosynthesis of an aliphatic halometabolite.

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Identification of polyketide synthase gene clusters in a phage P1-derived artificial chromosome library of a Philippine strain of Streptomyces sp. PCS3-D2

Asia Pacific Journal of Molecular Biology and Biotechnology ◽

10.35118/apjmbb.2019.027.2.08 ◽

2019 ◽

pp. 56-63

Author(s):

Aileen Bayot Custodio ◽

Edwin Plata Alcantara

Keyword(s):

Gene Cluster ◽

Polyketide Synthase ◽

Genetic Manipulation ◽

Gene Clusters ◽

Artificial Chromosome ◽

Restriction Enzyme Digestion ◽

Type I ◽

Putative Gene ◽

Streptomyces Sp ◽

Type Iii

A phage P1-derived artificial chromosome (PAC) library was constructed from genomic DNA of Streptomyces sp. PCS3-D2. Polymerase chain reaction (PCR) screening of the PAC library revealed two clones, PAC16D and P222O, which were positively identified to harbor polyketide synthase (PKS) Type I and PKS Type III gene clusters, respectively. Restriction enzyme digestion showed that PAC16D and PAC222O contained a 130 kb and a 140 kb insert, respectively. Results of sequencing and bioinformatics analyses revealed that PAC16D comprised of a full-length PKS type I bafilomycin gene cluster while PAC222O harbored truncated siderophore and putative gene clusters as well as a complete PKS III biosynthetic gene cluster. The PKS III gene cluster had three genes similar to alkyl resorcinol biosynthetic genes, however majority of the novel gene cluster had little similarity to known PKS Type III gene clusters. The successful cloning and identification of these gene clusters from Streptomyces sp. PCS3-D2 serve as the jump off point to further genetic manipulation in order to produce the insecticidal natural product in a heterologous host.

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