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Journal of Microbiology and... Jul 2023Type III polyketide synthase (PKS) found in bacteria is known as 1,3,6,8-tetrahydroxynaphthalene synthase (THNS). Microbial type III PKSs synthesize various compounds...
Type III polyketide synthase (PKS) found in bacteria is known as 1,3,6,8-tetrahydroxynaphthalene synthase (THNS). Microbial type III PKSs synthesize various compounds that possess crucial biological functions and significant pharmaceutical activities. Based on our sequence analysis, we have identified a putative type III polyketide synthase from sp. CS682 was named as ThnA. The role of ThnA, in sp. CS682 during the biosynthesis of 1,3,6,8 tetrahydroxynaphthalene (THN), which is the key intermediate of 1-(α-L-(2--methyl)-6-deoxymannopyranosyloxy)-3,6,8-trimethoxynaphthalene (IBR-3) was characterized. ThnA utilized five molecules of malonyl-CoA as a starter substrate to generate the polyketide 1,3,6,8-tetrahydroxynaphthalene, which could spontaneously be oxidized to the red flaviolin compound 2,5,7-trihydroxy-1,4-naphthoquinone. The amino acid sequence alignment of ThnA revealed similarities with a previously identified type III PKS and identified Cys, Phe, His, and Asn as four highly conserved active site amino acid residues, as found in other known polyketide synthases. In this study, we report the heterologous expression of the type III polyketide synthase in TK24 and the identification of THN production in a mutant strain. We also compared the transcription level of in TK24 and pIBR25-thnA and found that was only transcribed in the mutant.
Topics: Nocardia; Amino Acid Sequence; Naphthols; Polyketide Synthases
PubMed: 37254303
DOI: 10.4014/jmb.2303.03008 -
Microbiology Spectrum Aug 2022Eremophilanes are a large group of "sesquiterpenes" produced by plants and fungi, with more than 180 compounds being known in fungi alone. Many of these compounds are...
Eremophilanes are a large group of "sesquiterpenes" produced by plants and fungi, with more than 180 compounds being known in fungi alone. Many of these compounds are phytotoxic, antimicrobial, anticancer and immunomodulators, and hence are of great economic values. Acremeremophilanes A to O have earlier been reported in a marine isolate of sp. We report here the presence of Acremeremophilane I, G, K, N, and O, in a plant beneficial fungus , in a strain-specific manner. We also describe a novel, P strain-specific polyketide synthase (PKS) gene cluster in . This gene cluster, designated cluster, is absent in the genome of a Q strain of , and in other spp.; instead, a near identical cluster is present in the genome of the toxic mold Stachybotrys chartarum. Using gene knockout, we provide evidence that acremeremophilanes are biosynthesized via a polyketide route, and not via the mevalonate/terpene synthesis route as believed. We propose here that the 10-carbon skeleton is a product of polyketide synthase, to which a five-carbon isoprene unit is added by a prenyl transferase (PT), a gene for which is present next to the PKS gene in the genome. Based on this evidence, we propose that at least some of the eremophilanes classified in literature as sesquiterpenes (catalyzed by terpene cyclase) are actually meroterpenes (catalyzed by PKSs and PTs), and that the core moiety is not a sesquiterpene, but a hybrid polyketide/isoprene unit. The article contradicts the established fact that acremeremophilane metabolites produced by fungi are sesquiterpenes; instead, our findings suggest that at least some of these well-studied metabolites are of polyketide origin. Acremeremophilane metabolites are of medicinal significance, and the present findings have implications for the metabolic engineering of these metabolites and also their overproduction in microbial cell factories.
Topics: Carbon; Polycyclic Sesquiterpenes; Polyketide Synthases; Polyketides; Terpenes; Trichoderma
PubMed: 35938791
DOI: 10.1128/spectrum.01793-22 -
Scientific Reports May 2018To identify the species of butyrolactol-producing Streptomyces strain TP-A0882, whole genome-sequencing of three type strains in a close taxonomic relationship was...
To identify the species of butyrolactol-producing Streptomyces strain TP-A0882, whole genome-sequencing of three type strains in a close taxonomic relationship was performed. In silico DNA-DNA hybridization using the genome sequences suggested that Streptomyces sp. TP-A0882 is classified as Streptomyces diastaticus subsp. ardesiacus. Strain TP-A0882, S. diastaticus subsp. ardesiacus NBRC 15402, Streptomyces coelicoflavus NBRC 15399, and Streptomyces rubrogriseus NBRC 15455 harbor at least 14, 14, 10, and 12 biosynthetic gene clusters (BGCs), respectively, coding for nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs). All 14 gene clusters were shared by S. diastaticus subsp. ardesiacus strains TP-A0882 and NBRC 15402, while only four gene clusters were shared by the three distinct species. Although BGCs for bacteriocin, ectoine, indole, melanine, siderophores such as deferrioxamine, terpenes such as albaflavenone, hopene, carotenoid and geosmin are shared by the three species, many BGCs for secondary metabolites such as butyrolactone, lantipeptides, oligosaccharide, some terpenes are species-specific. These results indicate the possibility that strains belonging to the same species possess the same set of secondary metabolite-biosynthetic pathways, whereas strains belonging to distinct species have species-specific pathways, in addition to some common pathways, even if the strains are taxonomically close.
Topics: Bacterial Proteins; Genetic Variation; Peptide Synthases; Phylogeny; Polyketide Synthases; Streptomyces
PubMed: 29720592
DOI: 10.1038/s41598-018-24921-y -
The New Phytologist Oct 2022Plant polyketides are well-known for their crucial functions in plants and their importance in the context of human health. They are synthesized by type III polyketide...
Plant polyketides are well-known for their crucial functions in plants and their importance in the context of human health. They are synthesized by type III polyketide synthases (PKSs) and their final functional diversity is determined by post-PKS tailoring enzymes. Gerbera hybrida is rich in two defense-related polyketides: gerberin and parasorboside. Their synthesis is known to be initiated by GERBERA 2-PYRONE SYNTHASE 1 (G2PS1), but the polyketide reductases (PKRs) that determine their final structure have not yet been identified. We identified two PKR candidates in the pathway, GERBERA REDUCTASE 1 (GRED1) and GRED2. Gene expression and metabolite analysis of different gerbera tissues, cultivars, and transgenic gerbera plants, and in vitro enzyme assays, were performed for functional characterization of the enzymes. GRED1 and GRED2 catalyze the second reduction step in parasorboside biosynthesis. They reduce the proximal keto domain of the linear CoA bound intermediate before lactonization. We identified a crucial tailoring step in an important gerbera PKS pathway and show that plant polyketide biosynthesis shares processing strategies with fungi and bacteria. The two tailoring enzymes are recruited from the ancient sporopollenin biosynthetic pathway to a defense-related PKS pathway in gerbera. Our data provide an example of how plants recruit conserved genes to new functions in secondary metabolism that are important for environmental adaptation.
Topics: Asteraceae; Glucosides; Plants, Genetically Modified; Polyketide Synthases; Polyketides; Pyrones
PubMed: 35719102
DOI: 10.1111/nph.18328 -
Organic Letters Dec 2014Biochemical studies of purified and dissected fungal polyketide synthase and nonribosomal peptide synthetase (PKS-NRPS) hybrid enzymes involved in biosynthesis of...
Biochemical studies of purified and dissected fungal polyketide synthase and nonribosomal peptide synthetase (PKS-NRPS) hybrid enzymes involved in biosynthesis of pseurotin and aspyridone indicate that one α-methylation step during polyketide synthesis is a prerequisite and a key checkpoint for chain transfer between PKS and NRPS modules. In the absence of the resulting γ-methyl feature, the completed polyketide intermediate is offloaded as an α-pyrone instead of being aminoacylated by the NRPS domain. These examples illustrate that precisely timed tailoring domain activities play critical roles in the overall programming of the iterative PKS (and NRPS) functions.
Topics: Amino Acid Sequence; Biochemical Phenomena; Biological Factors; Fungi; Methylation; Molecular Structure; Peptide Synthases; Polyketide Synthases; Protein Engineering; Pyrones; Pyrrolidinones
PubMed: 25494235
DOI: 10.1021/ol503179v -
Biochemistry May 2014Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for... (Review)
Review
Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75-106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain-domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology.
Topics: Models, Chemical; Models, Molecular; Multienzyme Complexes; Polyketide Synthases; Protein Structure, Tertiary; Stereoisomerism
PubMed: 24779441
DOI: 10.1021/bi500290t -
Sequence, cloning, and analysis of the fluvirucin B1 polyketide synthase from Actinomadura vulgaris.ACS Synthetic Biology Nov 2013Fluvirucin B1 , produced by Actinomadura vulgaris, is a 14-membered macrolactam active against a variety of infectious fungi as well as influenza A. Despite considerable...
Fluvirucin B1 , produced by Actinomadura vulgaris, is a 14-membered macrolactam active against a variety of infectious fungi as well as influenza A. Despite considerable interest from the synthetic community, very little information is available regarding the biosynthetic origins of the fluvirucins. Herein, we report the identification and initial characterization of the fluvirucin B1 polyketide synthase and related enzymes. The cluster consists of five extender modules flanked by an N-terminal acyl carrier protein and C-terminal thioesterase domain. All but one of the synthase modules contain the full complement of tailoring domains (ketoreductase, dehydratase, and enoyl reductase) as determined by sequence homology with known polyketide synthases. Acitve site analyses of several key components of the cluster are performed to further verify that this gene cluster is associated with production of fluvirucin B1 . This work will both open doors toward a better understanding of macrolactam formation and provide an avenue to genetics-based diversification of fluvirucin structure.
Topics: Actinomycetales; Acyl Carrier Protein; Amino Acid Sequence; Bacterial Proteins; Cloning, Molecular; DNA, Bacterial; Hydro-Lyases; Lactams; Molecular Sequence Data; Multienzyme Complexes; Multigene Family; Polyketide Synthases; Sequence Alignment; Sequence Analysis, DNA
PubMed: 23654262
DOI: 10.1021/sb4000355 -
Acta Crystallographica. Section F,... Jun 2014CsyB from Aspergillus oryzae is a novel type III polyketide synthase that catalyzes the formation of csypyrone B1 [4-(3-acetyl-4-hydroxy-2-oxo-2H-pyran-6-yl)butyric...
CsyB from Aspergillus oryzae is a novel type III polyketide synthase that catalyzes the formation of csypyrone B1 [4-(3-acetyl-4-hydroxy-2-oxo-2H-pyran-6-yl)butyric acid] from fatty acyl-CoA, malonyl-CoA and acetoacetyl-CoA. Recombinant CsyB expressed in Escherichia coli was crystallized by the sitting-drop vapour-diffusion method. The crystals belonged to space P2₁, with unit-cell parameters a=70.0, b=104.8, c=73.5 Å, β=114.4°.
Topics: Aspergillus oryzae; Base Sequence; Crystallization; DNA Primers; Escherichia coli; Polyketide Synthases; Pyrones; Recombinant Proteins
PubMed: 24915080
DOI: 10.1107/S2053230X14008516 -
Applied and Environmental Microbiology Dec 2022Angucyclines are a family of structurally diverse, aromatic polyketides with some members that exhibit potent bioactivity. Angucyclines have also attracted considerable...
Angucyclines are a family of structurally diverse, aromatic polyketides with some members that exhibit potent bioactivity. Angucyclines have also attracted considerable attention due to the intriguing biosynthetic origins that underlie their structural complexity and diversity. Balmoralmycin (compound 1) represents a unique group of angucyclines that contain an angular benz[]anthracene tetracyclic system, a characteristic C-glycosidic bond-linked deoxy-sugar (d-olivose), and an unsaturated fatty acid chain. In this study, we identified a strain that produces balmoralmycin and seven biosynthetically related coproducts (compounds 2-8). Four of the coproducts (compounds 5-8) are novel compounds that feature a highly oxygenated or fragmented lactone ring, and three of them (compounds 3-5) exhibited cytotoxicity against the human pancreatic cancer cell line MIA PaCa-2 with IC values ranging from 0.9 to 1.2 μg/mL. Genome sequencing and CRISPR/dCas9-assisted gene knockdown led to the identification of the ~43 kb balmoralmycin biosynthetic gene cluster ( BGC). The BGC encodes a type II polyketide synthase (PKS) system for assembling the angucycline aglycone, six enzymes for generating the deoxysugar d-olivose, and a hybrid type II/III PKS system for synthesizing the 2,4-decadienoic acid chain. Based on the genetic and chemical information, we propose a mechanism for the biosynthesis of balmoralmycin and the shunt products. The chemical and genetic studies yielded insights into the biosynthetic origin of the structural diversity of angucyclines. Angucyclines are structurally diverse aromatic polyketides that have attracted considerable attention due to their potent bioactivity and intriguing biosynthetic origin. Balmoralmycin is a representative of a small family of angucyclines with unique structural features and an unknown biosynthetic origin. We report a newly isolated strain that produces balmoralmycin in a high fermentation titer as well as several structurally related shunt products. Based on the chemical and genetic information, a biosynthetic pathway that involves a type II polyketide synthase (PKS) system, cyclases/aromatases, oxidoreductases, and other ancillary enzymes was established. The elucidation of the balmoralmycin pathway enriches our understanding of how structural diversity is generated in angucyclines and opens the door for the production of balmoralmycin derivatives via pathway engineering.
Topics: Humans; Biosynthetic Pathways; Multigene Family; Polyketide Synthases; Polyketides; Streptomyces; Cell Line, Tumor
PubMed: 36350133
DOI: 10.1128/aem.01208-22 -
Marine Drugs Nov 2023The hadal biosphere is the most mysterious ecosystem on the planet, located in a unique and extreme environment on Earth. To adapt to extreme environmental conditions,...
The hadal biosphere is the most mysterious ecosystem on the planet, located in a unique and extreme environment on Earth. To adapt to extreme environmental conditions, hadal microorganisms evolve special strategies and metabolisms to survive and reproduce. However, the secondary metabolites of the hadal microorganisms are poorly understood. In this study, we focused on the isolation and characterization of hadal fungi, screening the potential strains with bioactive natural products. The isolates obtained were detected further for the polyketide synthase (PKS) genes. Two isolates of were picked up as the representatives, which had the potential to synthesize active natural products. The epigenetic modifiers were used for the two isolates to stimulate functional gene expression in hadal fungi under laboratory conditions. The results showed that the chemical epigenetic modifier, 5-Azacytidine (5-Aza), affected the phenotype, PKS gene expression, production of secondary metabolites, and antimicrobial activity of the hadal fungus . The influence of epigenetic modification on natural products was strongest when the concentration of 5-Aza was 50 μM. Furthermore, the modification of epigenetic agents on hadal fungi under high hydrostatic pressure (HHP) of 40 MPa displayed significant effects on PKS gene expression, and also activated the production of new compounds. Our study demonstrates the high biosynthetic potential of cultivable hadal fungi, but also provides evidence for the utility of chemical epigenetic modifiers on active natural products from hadal fungi, providing new ideas for the development and exploitation of microbial resources in extreme environments.
Topics: Ecosystem; Polyketide Synthases; Hydrostatic Pressure; Epigenesis, Genetic; Biological Products
PubMed: 37999409
DOI: 10.3390/md21110585