<i>O</i>- and <i>N</i>-Methylation in the Biosynthesis of Staurosporine

Yang, Shu‐Wei; Li, Lin; Cordell, Geoffrey A.; Wang, Ping; Corley, David G.

doi:10.1021/np990261q

Cited by 23 publications

(18 citation statements)

References 27 publications

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“…Streptomycetes produce a variety of bioactive polyketides, many of which contain methyl groups introduced via the action of N-and O-methyltransferases (16,66). PhzM most closely resembles O-methyltransferases from Streptomyces spp.…”

Section: Discussionmentioning

confidence: 99%

Functional Analysis of Genes for Biosynthesis of Pyocyanin and Phenazine-1-Carboxamide from Pseudomonas aeruginosa PAO1

et al. 2001

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Two seven-gene phenazine biosynthetic loci were cloned from Pseudomonas aeruginosa PAO1. The operons, designated phzA1B1C1D1E1F1G1 and phzA2B2C2D2E2F2G2, are homologous to previously studied phenazine biosynthetic operons from Pseudomonas fluorescens and Pseudomonas aureofaciens. Functional studies of phenazine-nonproducing strains of fluorescent pseudomonads indicated that each of the biosynthetic operons from P. aeruginosa is sufficient for production of a single compound, phenazine-1-carboxylic acid (PCA). Subsequent conversion of PCA to pyocyanin is mediated in P. aeruginosa by two novel phenazine-modifying genes, phzM and phzS, which encode putative phenazine-specific methyltransferase and flavin-containing monooxygenase, respectively. Expression of phzS alone in Escherichia coli or in enzymes, pyocyanin-nonproducing P. fluorescens resulted in conversion of PCA to 1-hydroxyphenazine. P. aeruginosa with insertionally inactivated phzM or phzS developed pyocyanin-deficient phenotypes. A third phenazine-modifying gene, phzH, which has a homologue in Pseudomonas chlororaphis, also was identified and was shown to control synthesis of phenazine-1-carboxamide from PCA in P. aeruginosa PAO1. Our results suggest that there is a complex pyocyanin biosynthetic pathway in P. aeruginosa consisting of two core loci responsible for synthesis of PCA and three additional genes encoding unique enzymes involved in the conversion of PCA to pyocyanin, 1-hydroxyphenazine, and phenazine-1-carboxamide.Phenazine compounds produced by fluorescent Pseudomonas species are biologically active metabolites that function in microbial competitiveness (37), the suppression of soilborne plant pathogens (1,11,55,56), and virulence in human and animal hosts (35).The most widely studied phenazine-producing fluorescent pseudomonad is P. aeruginosa, a gram-negative opportunistic pathogen of animals, insects, nematodes, and plants (30,33,35,46). In humans, P. aeruginosa infects immunocompromised, burned, or injured patients and can cause both acute and chronic lung disease. Strains of P. aeruginosa produce a variety of redox-active phenazine compounds, including pyocyanin, phenazine-1-carboxylic acid (PCA), 1-hydroxyphenazine (1-OH-PHZ), and phenazine-1-carboxamide (PCN) (7,52,57).From 90 to 95% of P. aeruginosa isolates produce pyocyanin (52), and the presence of high concentrations of pyocyanin in the sputum of cystic fibrosis patients has suggested that this compound plays a role in pulmonary tissue damage observed with chronic lung infections (64). This idea is supported by several recent studies which demonstrated that pyocyanin contributes in a variety of ways to the pathophysiological effects observed in airways infected by P. aeruginosa. Pyocyanin interferes with the regulation of ion transport, ciliary beat frequency, and mucus secretion in airway epithelial cells by altering the cytosolic concentration of calcium (15). It may interact with endothelium-derived relaxing factor or with nitric oxide (which plays a central role in the control ...

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Section: Discussionmentioning

confidence: 99%

Functional Analysis of Genes for Biosynthesis of Pyocyanin and Phenazine-1-Carboxamide from Pseudomonas aeruginosa PAO1

et al. 2001

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“…7) Biosynthetic studies done with isotope-labeled precursors indicated that the indolocarbazole structure is derived from two molecules of tryptophan, probably via indolepyruvic acid as an intermediate. [8][9][10] Ohuchi et al reported cloning of the ngt gene encoding an Nglycosyltransferase from L. aerocolonigenes ATCC 39243. 5) The Ngt protein converted an indolocarbazole, J-104303, to its N-glucoside, indicating that ngt is responsible for N-glycosylation in rebeccamycin biosynthesis.…”

Section: Characterization Of the Biosynthetic Gene Cluster Ofmentioning

confidence: 99%

Characterization of the Biosynthetic Gene Cluster of Rebeccamycin fromLechevalieria aerocolonigenesATCC 39243

Onaka¹,

Taniguchi²,

Igarashi³

et al. 2003

Bioscience, Biotechnology, and Biochemistry

112

124

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The biosynthetic gene cluster for rebeccamycin, an indolocarbazole antibiotic, from Lechevalieria aerocolonigenes ATCC 39243 has 11 ORFs. To clarify their functions, mutants with rebG, rebD, rebC, rebP, rebM, rebR, rebH, rebT, or orfD2 disrupted were constructed, and the gene products were examined. rebP disruptants produced 11,11'-dichlorochromopyrrolic acid, found to be a biosynthetic intermediate by a bioconversion experiment. Other genes encoded N-glycosyltransferase (rebG), monooxygenase (rebC), methyltransferase (rebM), a transcriptional activator (rebR), and halogenase (rebH). rebT disruptants produced rebeccamycin as much as the wild strain, so rebT was probably not involved in rebeccamycin production. Biosynthetic genes of staurosporine, an another indolocarbazole antibiotic, were cloned from Streptomyces sp. TP-A0274. staO, staD, and staP were similar to rebO, rebD, and rebP, respectively, all of which are responsible for indolocarbazole biosynthesis, But a rebC homolog, encoding a putative enzyme oxidizing the C-7 site of pyrrole rings, was not found in the staurosporine biosynthetic gene cluster. These results suggest that indolocarbazole is constructed by oxidative decarboxylation of chromopyrrolic acid (11,11'-dichlorochromopyrrolic acid in rebeccamycin) generated from two molecules of tryptophan by coupling and that the oxidation state at the C-7 position depends on the additional enzyme(s) encoded by the biosynthetic genes.

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“…PhzM is a predicted 36-kDa protein similar to enzymes involved in the methylation of polyketide antibiotics by Streptomyces spp. (60). Though the 1.8-Å crystal structure of PhzM from P. aeruginosa was determined by single-wavelength anomalous dispersion (40), little is known about the features of phzM gene expression and regulation in pseudomonads.…”

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confidence: 99%

Temperature-Dependent Expression of phzM and Its Regulatory Genes lasI and ptsP in Rhizosphere Isolate Pseudomonas sp. Strain M18

Huang

Zhang

et al. 2009

Appl Environ Microbiol

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Pseudomonas sp. strain M18, an effective biological control agent isolated from the melon rhizosphere, has a genetic background similar to that of the opportunistic human pathogen Pseudomonas aeruginosa PAO1. However, the predominant phenazine produced by strain M18 is phenazine-1-carboxylic acid (PCA) rather than pyocyanin (PYO); the quantitative ratio of PCA to PYO is 105 to 1 at 28°C in strain M18, while the ratio is 1 to 2 at 37°C in strain PAO1. We first provided evidence that the differential production of the two phenazines in strains M18 and PAO1 is related to the temperature-dependent and strain-specific expression patterns of phzM, a gene involved in the conversion of PCA to PYO. Transcriptional levels of phzM were measured by quantitative real-time PCR, and the activities of both transcriptional and translational phzM-lacZ fusions were determined in strains M18 and PAO1, respectively. Using lasI::Gm and ptsP::Gm inactivation M18 mutants, we further show that expression of the phzM gene is positively regulated by the quorum-sensing protein LasI and negatively regulated by the phosphoenolpyruvate phosphotransferase protein PtsP. Surprisingly, the lasI and ptsP regulatory genes were also expressed in a temperature-dependent and strain-specific manner. The differential production of the phenazines PCA and PYO by strains M18 and PAO1 may be a consequence of selective pressure imposed on P. aeruginosa PAO1 and its relative M18 in the two different niches over a long evolutionary process.Phenazines, known for over 150 years, are a group of the nitrogen-containing secondary metabolites synthesized mainly by Pseudomonas spp. and a few other bacterial strains. Advances within the past 2 decades have provided significant new insights into the genetics, biochemistry, and regulation of phenazine synthesis, as well as the mode of action and functional roles of these compounds in various environments (33,44). Despite the fact that the phenazine biosynthesis locus is highly conserved among various Pseudomonas spp., individual strains differ in the range of phenazine compounds they produce and the relative amounts of the possible phenazines are strongly influenced by growth conditions (7,30,34,61). The crucial roles of phenazine-1-carboxylic acid (PCA) in plant root disease suppression has been well documented in studies with several biological control strains, such as Pseudomonas fluorescens 2-79, P. chlororaphis 30-84, and P. chlororaphis PCL1391, where PCA can be converted into phenazine-1-carboxamide (PCN) (6,33,35,37,50). Nevertheless, PCA is considered a predominant phenazine (44) in these strains, and its secretion mainly contributes to the biocontrol activity against various fungal phytopathogens such as Gaeumannomyces graminis var. tritici (41,49,50). Chromosomal insertion of genes involved in the PCA biosynthetic pathway enhances the efficacy of damping-off disease control by P. fluorescens. The phenazine-deficient strains P. fluorescens 2-79 and P. chlororaphis 30-84 have reduced survival rates and a dimini...

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O- and N-Methylation in the Biosynthesis of Staurosporine

Cited by 23 publications

References 27 publications

Functional Analysis of Genes for Biosynthesis of Pyocyanin and Phenazine-1-Carboxamide from Pseudomonas aeruginosa PAO1

Functional Analysis of Genes for Biosynthesis of Pyocyanin and Phenazine-1-Carboxamide from Pseudomonas aeruginosa PAO1

Characterization of the Biosynthetic Gene Cluster of Rebeccamycin fromLechevalieria aerocolonigenesATCC 39243

Temperature-Dependent Expression of phzM and Its Regulatory Genes lasI and ptsP in Rhizosphere Isolate Pseudomonas sp. Strain M18

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