scholarly journals Molecular Basis and Phylogenetic Implications of Deoxycylindrospermopsin Biosynthesis in the Cyanobacterium Raphidiopsis curvata

2012 ◽  
Vol 78 (7) ◽  
pp. 2256-2263 ◽  
Author(s):  
Yongguang Jiang ◽  
Peng Xiao ◽  
Gongliang Yu ◽  
Tomoharu Sano ◽  
Qianqian Pan ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Phylogenetic Trees ◽  
Purifying Selection ◽  
Neutral Evolution ◽  
Cylindrospermopsis Raciborskii ◽  
Insertion Mutation ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Cell Extracts ◽  
Phylogenetic Implications

ABSTRACTNew insights into the distribution and biochemistry of the cyanotoxin cylindrospermopsin (CYN) have been provided by the recent determination of its biosynthesis gene cluster (cyr) in several cyanobacterial species.Raphidiopsis curvataCHAB1150 isolated from China was analyzed for CYN analogues. Only 7-deoxy-CYN was detected in the cell extracts. Thecyrgene cluster ofR. curvataCHAB1150 was sequenced, and thecyrgenes of this strain were found to have extremely high similarities (96% to 100%) to those from other nostocalean species. These species includeCylindrospermopsis raciborskiiAWT205,Aphanizomenonsp. strain 10E6, andAphanizomenon ovalisporumILC-146. Insertion mutation was identified within thecyrIgene, and transcripts ofcyrIand another functional genecyrJwere detected inR. curvataCHAB1150. General congruence between the phylogenetic trees based on bothcyrand 16Srrnwas displayed. Neutral evolution was found on the whole sequences of thecyrgenes, and 0 to 89 negative selected codons were detected in each gene. Therefore, the function of CyrI is to catalyze the oxygenation of 7-deoxy-CYN in CYN biosynthesis. The transcripts of the mutatedcyrIgene may result from polycistronic transcription. The high conservation of thecyrgenes may be ascribed to purifying selection and horizontal gene transfer.

scholarly journals Overproduction of Ristomycin A by Activation of a Silent Gene Cluster in Amycolatopsis japonicum MG417-CF17

2014 ◽  
Vol 58 (10) ◽  
pp. 6185-6196 ◽  
Author(s):  
Marius Spohn ◽  
Norbert Kirchner ◽  
Andreas Kulik ◽  
Angelika Jochim ◽  
Felix Wolf ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Gene Cluster ◽  
High Performance ◽  
Pathogenic Bacteria ◽  
Genome Mining ◽  
Gene Clusters ◽  
Von Willebrand Disease ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Bioinformatic Tools

ABSTRACTThe emergence of antibiotic-resistant pathogenic bacteria within the last decades is one reason for the urgent need for new antibacterial agents. A strategy to discover new anti-infective compounds is the evaluation of the genetic capacity of secondary metabolite producers and the activation of cryptic gene clusters (genome mining). One genus known for its potential to synthesize medically important products isAmycolatopsis. However,Amycolatopsis japonicumdoes not produce an antibiotic under standard laboratory conditions. In contrast to mostAmycolatopsisstrains,A. japonicumis genetically tractable with different methods. In order to activate a possible silent glycopeptide cluster, we introduced a gene encoding the transcriptional activator of balhimycin biosynthesis, thebbrgene fromAmycolatopsis balhimycina(bbrAba), intoA. japonicum. This resulted in the production of an antibiotically active compound. Following whole-genome sequencing ofA. japonicum, 29 cryptic gene clusters were identified by genome mining. One of these gene clusters is a putative glycopeptide biosynthesis gene cluster. Using bioinformatic tools, ristomycin (syn. ristocetin), a type III glycopeptide, which has antibacterial activity and which is used for the diagnosis of von Willebrand disease and Bernard-Soulier syndrome, was deduced as a possible product of the gene cluster. Chemical analyses by high-performance liquid chromatography and mass spectrometry (HPLC-MS), tandem mass spectrometry (MS/MS), and nuclear magnetic resonance (NMR) spectroscopy confirmed thein silicoprediction that the recombinantA. japonicum/pRM4-bbrAbasynthesizes ristomycin A.

scholarly journals Clostridium acidurici Electron-Bifurcating Formate Dehydrogenase

2013 ◽  
Vol 79 (19) ◽  
pp. 6176-6179 ◽  
Author(s):  
Shuning Wang ◽  
Haiyan Huang ◽  
Jörg Kahnt ◽  
Rudolf K. Thauer
Keyword(s):  
Uric Acid ◽  
Gene Cluster ◽  
Formate Dehydrogenase ◽  
Specific Activity ◽  
Enzyme Complex ◽  
Content Type ◽  
Cell Extracts

ABSTRACTCell extracts of uric acid-grownClostridium aciduricicatalyzed the coupled reduction of NAD+and ferredoxin with formate at a specific activity of 1.3 U/mg. The enzyme complex catalyzing the electron-bifurcating reaction was purified 130-fold and found to be composed of four subunits encoded by the gene clusterhylCBA-fdhF2.

scholarly journals Characterization of the Amicetin Biosynthesis Gene Cluster from Streptomyces vinaceusdrappus NRRL 2363 Implicates Two Alternative Strategies for Amide Bond Formation

2012 ◽  
Vol 78 (7) ◽  
pp. 2393-2401 ◽  
Author(s):  
Gaiyun Zhang ◽  
Haibo Zhang ◽  
Sumei Li ◽  
Ji Xiao ◽  
Guangtao Zhang ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Acyl Carrier Protein ◽  
Bond Formation ◽  
Antiviral Agent ◽  
Amide Bond ◽  
Biosynthesis Gene Cluster ◽  
Alternative Strategies ◽  
Content Type ◽  
Amide Bond Formation ◽  
Biosynthesis Gene

ABSTRACTAmicetin, an antibacterial and antiviral agent, belongs to a group of disaccharide nucleoside antibiotics featuring an α-(1→4)-glycoside bond in the disaccharide moiety. In this study, the amicetin biosynthesis gene cluster was cloned fromStreptomyces vinaceusdrappusNRRL 2363 and localized on a 37-kb contiguous DNA region. Heterologous expression of the amicetin biosynthesis gene cluster inStreptomyces lividansTK64 resulted in the production of amicetin and its analogues, thereby confirming the identity of theamigene cluster.In silicosequence analysis revealed that 21 genes were putatively involved in amicetin biosynthesis, including 3 for regulation and transportation, 10 for disaccharide biosynthesis, and 8 for the formation of the amicetin skeleton by the linkage of cytosine,p-aminobenzoic acid (PABA), and the terminal (+)-α-methylserine moieties. The inactivation of the benzoate coenzyme A (benzoate-CoA) ligase geneamiLand theN-acetyltransferase geneamiFled to two mutants that accumulated the same two compounds, cytosamine and 4-acetamido-3-hydroxybenzoic acid. These data indicated that AmiF functioned as an amide synthethase to link cytosine and PABA. The inactivation ofamiR, encoding an acyl-CoA-acyl carrier protein transacylase, resulted in the production of plicacetin and norplicacetin, indicating AmiR to be responsible for attachment of the terminal methylserine moiety to form another amide bond. These findings implicated two alternative strategies for amide bond formation in amicetin biosynthesis.

scholarly journals Cloning and Characterization of the Polyether Salinomycin Biosynthesis Gene Cluster of Streptomyces albus XM211

2011 ◽  
Vol 78 (4) ◽  
pp. 994-1003 ◽  
Author(s):  
Chunyan Jiang ◽  
Hougen Wang ◽  
Qianjin Kang ◽  
Jing Liu ◽  
Linquan Bai
Keyword(s):  
Gene Cluster ◽  
Animal Husbandry ◽  
Oxidative Cyclization ◽  
Degenerate Primers ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Feeding Experiments ◽  
Streptomyces Albus ◽  
Polyketide Chain

ABSTRACTSalinomycin is widely used in animal husbandry as a food additive due to its antibacterial and anticoccidial activities. However, its biosynthesis had only been studied by feeding experiments with isotope-labeled precursors. A strategy with degenerate primers based on the polyether-specific epoxidase sequences was successfully developed to clone the salinomycin gene cluster. Using this strategy, a putative epoxidase gene,slnC, was cloned from the salinomycin producerStreptomyces albusXM211. The targeted replacement ofslnCand subsequenttrans-complementation proved its involvement in salinomycin biosynthesis. A 127-kb DNA region containingslnCwas sequenced, including genes for polyketide assembly and release, oxidative cyclization, modification, export, and regulation. In order to gain insight into the salinomycin biosynthesis mechanism, 13 gene replacements and deletions were conducted. IncludingslnC, 7 genes were identified as essential for salinomycin biosynthesis and putatively responsible for polyketide chain release, oxidative cyclization, modification, and regulation. Moreover, 6 genes were found to be relevant to salinomycin biosynthesis and possibly involved in precursor supply, removal of aberrant extender units, and regulation. Sequence analysis and a series of gene replacements suggest a proposed pathway for the biosynthesis of salinomycin. The information presented here expands the understanding of polyether biosynthesis mechanisms and paves the way for targeted engineering of salinomycin activity and productivity.

scholarly journals Molecular Basis for Mycophenolic Acid Biosynthesis in Penicillium brevicompactum

2011 ◽  
Vol 77 (9) ◽  
pp. 3035-3043 ◽  
Author(s):  
Torsten Bak Regueira ◽  
Kanchana Rueksomtawin Kildegaard ◽  
Bjarne Gram Hansen ◽  
Uffe H. Mortensen ◽  
Christian Hertweck ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Mycophenolic Acid ◽  
Molecular Basis ◽  
Polyketide Synthase ◽  
Functional Characterization ◽  
Gene Encoding ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Polyketide Synthase Gene ◽  
Penicillium Brevicompactum

ABSTRACTMycophenolic acid (MPA) is the active ingredient in the increasingly important immunosuppressive pharmaceuticals CellCept (Roche) and Myfortic (Novartis). Despite the long history of MPA, the molecular basis for its biosynthesis has remained enigmatic. Here we report the discovery of a polyketide synthase (PKS), MpaC, which we successfully characterized and identified as responsible for MPA production inPenicillium brevicompactum. mpaCresides in what most likely is a 25-kb gene cluster in the genome ofPenicillium brevicompactum. The gene cluster was successfully localized by targeting putative resistance genes, in this case an additional copy of the gene encoding IMP dehydrogenase (IMPDH). We report the cloning, sequencing, and the functional characterization of the MPA biosynthesis gene cluster by deletion of the polyketide synthase genempaCofP. brevicompactumand bioinformatic analyses. As expected, the gene deletion completely abolished MPA production as well as production of several other metabolites derived from the MPA biosynthesis pathway ofP. brevicompactum. Our work sets the stage for engineering the production of MPA and analogues through metabolic engineering.

scholarly journals Whole-Genome Sequencing of the Fungus Penicillium citrinum Reveals the Biosynthesis Gene Cluster for the Mycotoxin Citrinin

2019 ◽  
Vol 8 (4) ◽  
Author(s):  
Markus Schmidt-Heydt ◽  
Dominic Stoll ◽  
Rolf Geisen
Keyword(s):  
Antibiotic Resistance ◽  
Whole Genome Sequencing ◽  
Gene Cluster ◽  
Genome Sequencing ◽  
Penicillium Citrinum ◽  
Whole Genome ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Biosynthesis Gene

Penicillium citrinum is a food-contaminating ascomycete that consistently produces large amounts of the mycotoxin citrinin. Citrinin exhibits, besides its toxicity, antibiotic effects and thus potentially forces antibiotic resistance.

scholarly journals Biosynthetic Intermediate Analysis and Functional Homology Reveal a Saxitoxin Gene Cluster in Cyanobacteria

2008 ◽  
Vol 74 (13) ◽  
pp. 4044-4053 ◽  
Author(s):  
Ralf Kellmann ◽  
Troco Kaan Mihali ◽  
Young Jae Jeon ◽  
Russell Pickford ◽  
Francesco Pomati ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Pathway ◽  
Condensation Reaction ◽  
Comparative Sequence Analysis ◽  
Mass Spectrometry Analysis ◽  
Cylindrospermopsis Raciborskii ◽  
Shellfish Poisoning ◽  
Biosynthesis Gene Cluster ◽  
Spectrometry Analysis ◽  
Psp Toxins

ABSTRACT Saxitoxin (STX) and its analogues cause the paralytic shellfish poisoning (PSP) syndrome, which afflicts human health and impacts coastal shellfish economies worldwide. PSP toxins are unique alkaloids, being produced by both prokaryotes and eukaryotes. Here we describe a candidate PSP toxin biosynthesis gene cluster (sxt) from Cylindrospermopsis raciborskii T3. The saxitoxin biosynthetic pathway is encoded by more than 35 kb, and comparative sequence analysis assigns 30 catalytic functions to 26 proteins. STX biosynthesis is initiated with arginine, S-adenosylmethionine, and acetate by a new type of polyketide synthase, which can putatively perform a methylation of acetate, and a Claisen condensation reaction between propionate and arginine. Further steps involve enzymes catalyzing three heterocyclizations and various tailoring reactions that result in the numerous isoforms of saxitoxin. In the absence of a gene transfer system in these microorganisms, we have revised the description of the known STX biosynthetic pathway, with in silico functional inferences based on sxt open reading frames combined with liquid chromatography-tandem mass spectrometry analysis of the biosynthetic intermediates. Our results indicate the evolutionary origin for the production of PSP toxins in an ancestral cyanobacterium with genetic contributions from diverse phylogenetic lineages of bacteria and provide a quantum addition to the catalytic collective available for future combinatorial biosyntheses. The distribution of these genes also supports the idea of the involvement of this gene cluster in STX production in various cyanobacteria.

scholarly journals High-Throughput Screening for Streptomyces Antibiotic Biosynthesis Activators

2012 ◽  
Vol 78 (12) ◽  
pp. 4526-4528 ◽  
Author(s):  
Li Chen ◽  
Yemin Wang ◽  
Hang Guo ◽  
Min Xu ◽  
Zixin Deng ◽  
...  
Keyword(s):  
Gene Cluster ◽  
High Throughput ◽  
High Throughput Screening ◽  
Streptomyces Clavuligerus ◽  
Cosmid Library ◽  
Screening Procedure ◽  
Antibiotic Biosynthesis ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Biosynthesis Gene

ABSTRACTA genomic cosmid library ofStreptomyces clavuligeruswas constructed and transferred efficiently by conjugation toStreptomyces lividans, and 12 distinct groups of overlapping cosmid clones that activated the silent actinorhodin biosynthesis gene cluster were identified. This generally applicable high-throughput screening procedure greatly facilitates the identification of antibiotic biosynthesis activators.

scholarly journals The Swinholide Biosynthesis Gene Cluster from a Terrestrial Cyanobacterium, Nostoc sp. Strain UHCC 0450

2017 ◽  
Vol 84 (3) ◽  
Author(s):  
Anu Humisto ◽  
Jouni Jokela ◽  
Liwei Liu ◽  
Matti Wahlsten ◽  
Hao Wang ◽  
...  
Keyword(s):  
Natural Products ◽  
Actin Cytoskeleton ◽  
Gene Cluster ◽  
Marine Sponge ◽  
Gene Clusters ◽  
Fold Axis ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Biosynthesis Gene ◽  
Axis Of Symmetry

ABSTRACT Swinholides are 42-carbon ring polyketides with a 2-fold axis of symmetry. They are potent cytotoxins that disrupt the actin cytoskeleton. Swinholides were discovered from the marine sponge Theonella sp. and were long suspected to be produced by symbiotic bacteria. Misakinolide, a structural variant of swinholide, was recently demonstrated to be the product of a symbiotic heterotrophic proteobacterium. Here, we report the production of swinholide A by an axenic strain of the terrestrial cyanobacterium Nostoc sp. strain UHCC 0450. We located the 85-kb trans -AT polyketide synthase (PKS) swinholide biosynthesis gene cluster from a draft genome of Nostoc sp. UHCC 0450. The swinholide and misakinolide biosynthesis gene clusters share an almost identical order of catalytic domains, with 85% nucleotide sequence identity, and they group together in phylogenetic analysis. Our results resolve speculation around the true producer of swinholides and demonstrate that bacteria belonging to two distantly related phyla both produce structural variants of the same natural product. In addition, we described a biosynthesis cluster from Anabaena sp. strain UHCC 0451 for the synthesis of the cytotoxic and antifungal scytophycin. All of these biosynthesis gene clusters were closely related to each other and created a group of cytotoxic macrolide compounds produced by trans -AT PKSs of cyanobacteria and proteobacteria. IMPORTANCE Many of the drugs in use today originate from natural products. New candidate compounds for drug development are needed due to increased drug resistance. An increased knowledge of the biosynthesis of bioactive compounds can be used to aid chemical synthesis to produce novel drugs. Here, we show that a terrestrial axenic culture of Nostoc cyanobacterium produces swinholides, which have been previously found only from marine sponge or samples related to them. Swinholides are polyketides with a 2-fold axis of symmetry, and they are potent cytotoxins that disrupt the actin cytoskeleton. We describe the biosynthesis gene clusters of swinholide from Nostoc cyanobacteria, as well as the related cytotoxic and antifungal scytophycin from Anabaena cyanobacteria, and we study the evolution of their trans -AT polyketide synthases. Interestingly, swinholide is closely related to misakinolide produced by a symbiotic heterotrophic proteobacterium, demonstrating that bacteria belonging to two distantly related phyla and different habitats can produce similar natural products.

scholarly journals Identification of a Polyketide Synthase Gene Responsible for Ascochitine Biosynthesis in Ascochyta fabae and Its Abrogation in Sister Taxa

mSphere ◽  
2019 ◽  
Vol 4 (5) ◽  
Author(s):  
Wonyong Kim ◽  
Judith Lichtenzveig ◽  
Robert A. Syme ◽  
Angela H. Williams ◽  
Tobin L. Peever ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Secondary Metabolite ◽  
Polyketide Synthase ◽  
Animal Health ◽  
Gene Clusters ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
Pathogenicity Factor ◽  
Biosynthesis Gene ◽  
Ascochyta Fabae

ABSTRACT The polyketide-derived secondary metabolite ascochitine is produced by species in the Didymellaceae family, including but not restricted to Ascochyta species pathogens of cool-season food legumes. Ascochitine is structurally similar to the well-known mycotoxin citrinin and exhibits broad-spectrum phytotoxicity and antimicrobial activities. Here, we identified a polyketide synthase (PKS) gene (denoted pksAC) responsible for ascochitine production in the filamentous fungus Ascochyta fabae. Deletion of the pksAC prevented production of ascochitine and its derivative ascochital in A. fabae. The putative ascochitine biosynthesis gene cluster comprises 11 genes that have undergone rearrangement and gain-and-loss events relative to the citrinin biosynthesis gene cluster in Monascus ruber. Interestingly, we also identified pksAC homologs in two recently diverged species, A. lentis and A. lentis var. lathyri, that are sister taxa closely related to ascochitine producers such as A. fabae and A. viciae-villosae. However, nonsense mutations have been independently introduced in coding sequences of the pksAC homologs of A. lentis and A. lentis var. lathyri that resulted in loss of ascochitine production. Despite its reported phytotoxicity, ascochitine was not a pathogenicity factor in A. fabae infection and colonization of faba bean (Vicia faba L.). Ascochitine was mainly produced from mature hyphae at the site of pycnidial formation, suggesting a possible protective role of the compound against other microbial competitors in nature. This report highlights the evolution of gene clusters harnessing the structural diversity of polyketides and a mechanism with the potential to alter secondary metabolite profiles via single nucleotide polymorphisms in closely related fungal species. IMPORTANCE Fungi produce a diverse array of secondary metabolites, many of which are of pharmacological importance whereas many others are noted for mycotoxins, such as aflatoxin and citrinin, that can threaten human and animal health. The polyketide-derived compound ascochitine, which is structurally similar to citrinin mycotoxin, has been considered to be important for pathogenicity of legume-associated Ascochyta species. Here, we identified the ascochitine polyketide synthase (PKS) gene in Ascochyta fabae and its neighboring genes that may be involved in ascochitine biosynthesis. Interestingly, the ascochitine PKS genes in other legume-associated Ascochyta species have been mutated, encoding truncated PKSs. This indicated that point mutations may have contributed to genetic diversity for secondary metabolite production in these fungi. We also demonstrated that ascochitine is not a pathogenicity factor in A. fabae. The antifungal activities and production of ascochitine during sporulation suggested that it may play a role in competition with other saprobic fungi in nature.

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