Fur from Microcystis aeruginosa binds in vitro promoter regions of the microcystin biosynthesis gene cluster

2006 ◽  
Vol 67 (9) ◽  
pp. 876-881 ◽  
Author(s):  
Beatriz Martin-Luna ◽  
Emma Sevilla ◽  
José A. Hernandez ◽  
M. Teresa Bes ◽  
Maria F. Fillat ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Microcystis Aeruginosa ◽  
Promoter Regions ◽  
Biosynthesis Gene Cluster ◽  
Biosynthesis Gene

scholarly journals Characterization of the 2-Hydroxy-acid Dehydrogenase McyI, Encoded within the Microcystin Biosynthesis Gene Cluster of Microcystis aeruginosa PCC7806

2006 ◽  
Vol 282 (7) ◽  
pp. 4681-4692 ◽  
Author(s):  
Leanne A. Pearson ◽  
Kevin D. Barrow ◽  
Brett A. Neilan
Keyword(s):  
Gene Cluster ◽  
Microcystis Aeruginosa ◽  
Hydroxy Acid ◽  
Biosynthesis Gene Cluster ◽  
Glutamate Mutase ◽  
Acid Dehydrogenase ◽  
Methyl Aspartate ◽  
Catalyzed Reactions

The cyanobacterium Microcystis aeruginosa is widely known for its production of the potent hepatotoxin microcystin. This cyclic heptapeptide is synthesized non-ribosomally by the thio-template function of a large modular enzyme complex encoded within the 55-kb microcystin synthetase gene (mcy) cluster. The mcy gene cluster also encodes several stand-alone enzymes, putatively involved in the tailoring and export of microcystin. This study describes the characterization of the 2-hydroxy-acid dehydrogenase McyI, putatively involved in the production of d-methyl aspartate at position 3 within the microcystin cyclic structure. A combination of bioinformatics, molecular, and biochemical techniques was used to elucidate the structure, function, regulation, and evolution of this unique enzyme. The recombinant McyI enzyme was overexpressed in Escherichia coli and enzymatically characterized. The hypothesized native activity of McyI, the interconversion of 3-methyl malate to 3-methyl oxalacetate, was demonstrated using an in vitro spectrophotometric assay. The enzyme was also able to reduce α-ketoglutarate to 2-hydroxyglutarate and to catalyze the interconversion of malate and oxalacetate. Although NADP(H) was the preferred cofactor of the McyI-catalyzed reactions, NAD(H) could also be utilized, although rates of catalysis were significantly lower. The combined results of this study suggest that hepatotoxic cyanobacteria such as M. aeruginosa PCC7806 are capable of producing methyl aspartate via a novel glutamate mutase-independent pathway, in which McyI plays a pivotal role.

scholarly journals Light-Induced Carotenogenesis in Streptomyces coelicolor A3(2): Identification of an Extracytoplasmic Function Sigma Factor That Directs Photodependent Transcription of the Carotenoid Biosynthesis Gene Cluster

2005 ◽  
Vol 187 (5) ◽  
pp. 1825-1832 ◽  
Author(s):  
Hideaki Takano ◽  
Saemi Obitsu ◽  
Teruhiko Beppu ◽  
Kenji Ueda
Keyword(s):  
Gene Expression ◽  
Gene Cluster ◽  
Sigma Factor ◽  
Streptomyces Coelicolor ◽  
Regulatory Region ◽  
Carotenoid Biosynthesis ◽  
Biosynthesis Gene Cluster ◽  
Biosynthesis Gene ◽  
Extracytoplasmic Function Sigma Factor

ABSTRACT Carotenoids are produced by a variety of organisms, but the mechanisms that regulate gene expression leading to carotenoid biosynthesis have been characterized for only a few organisms. In this study, we found that Streptomyces coelicolor A3(2), a gram-positive filamentous bacterium, produces carotenoids under blue light induction. The carotenoid fraction isolated from the cell extract contained multiple compounds, including isorenieratene and β-carotene. The carotenoid biosynthesis gene cluster of S. coelicolor consists of two convergent operons, crtEIBV and crtYTU, as previously shown for Streptomyces griseus. The crtEIBV null mutant completely lost its ability to produce carotenoids. The crt gene cluster is flanked by a regulatory region that consists of two divergent operons, litRQ and litSAB. The lit (light-induced transcription) genes encode a MerR-type transcriptional regulator (LitR), a possible oxidoreductase (LitQ), an extracytoplasmic function sigma factor (σLitS), a putative lipoprotein (LitA), and a putative anti-sigma factor (LitB). S1 protection assay revealed that the promoters preceding crtE (PcrtE), crtY (PcrtY), litR (PlitR), and litS (PlitS) are activated upon illumination. A litS mutant lost both the ability to produce carotenoids and the activities of PcrtE, PcrtY, and PlitS, which suggested that σLitS directs light-induced transcription from these promoters. An RNA polymerase holocomplex containing purified σLitS recombinant protein generated specific PcrtE and PcrtY transcripts in an in vitro runoff transcriptional assay. A litR mutant that had an insertion of the kanamycin resistance gene was defective both in the ability to produce carotenoids and in all of the light-dependent promoter activities. Overexpression of litS resulted in constitutive carotenoid production in both the wild type and the litR mutant. These results indicate that σLitS acts as a light-induced sigma factor that directs transcription of the crt biosynthesis gene cluster, whose activity is controlled by an unknown LitR function. This is the first report to describe light-inducible gene expression in Streptomyces.

scholarly journals Distribution and conservation of known secondary metabolite biosynthesis gene clusters in the genomes of geographically diverse Microcystis aeruginosa strains

2020 ◽  
Vol 71 (5) ◽  
pp. 701 ◽  
Author(s):  
Leanne A. Pearson ◽  
Nicholas D. Crosbie ◽  
Brett A. Neilan
Keyword(s):  
Gene Cluster ◽  
Secondary Metabolite ◽  
Microcystis Aeruginosa ◽  
Gene Clusters ◽  
Biologically Active ◽  
Biosynthesis Gene Cluster ◽  
Phosphatase Inhibitors ◽  
Biosynthesis Gene ◽  
Screening Study ◽  
Non Ribosomal Peptides

The cyanobacterium Microcystis aeruginosa has been linked to toxic blooms worldwide. In addition to producing hepatotoxic microcystins, many strains are capable of synthesising a variety of biologically active compounds, including protease and phosphatase inhibitors, which may affect aquatic ecosystems and pose a risk to their use. This study explored the distribution, composition and conservation of known secondary metabolite (SM) biosynthesis gene clusters in the genomes of 27 M. aeruginosa strains isolated from six different Köppen–Geiger climates. Our analysis identified gene clusters with significant homology to nine SM biosynthesis gene clusters spanning four different compound classes: non-ribosomal peptides, hybrid polyketide–non-ribosomal peptides, cyanobactins and microviridins. The aeruginosin, microviridin, cyanopeptolin and microcystin biosynthesis gene clusters were the most frequently observed, but hybrid polyketide–non-ribosomal peptide biosynthesis clusters were the most common class overall. Although some biogeographic relationships were observed, taxonomic markers and geography were not reliable indicators of SM biosynthesis cluster distribution, possibly due to previous genetic deletions or horizontal gene transfer events. The only cyanotoxin biosynthesis gene cluster identified in our screening study was the microcystin synthetase (mcy) gene cluster, suggesting that the production of non-microcystin cyanotoxins by this taxon, such as anatoxin-a or paralytic shellfish poison analogues, is either absent or rare.

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