Gene arrangement, functional annotation and domain stru

A hybrid non-ribosomal peptide/polyketide synthetase containing fatty-acyl ligase (FAAL) synthesizes the β-amino fatty acid lipopeptides puwainaphycins in the Cyanobacterium Cylindrospermum alatosporum. Mareš J, Hájek J, Urajová P, Kopecký J, Hrouzek P - PLoS ONE (2014) Bottom Line: Bioinformatics analysis enabled sequential prediction of puwainaphycin biosynthesis; this process is initiated by the activation of a fatty acid residue via fatty acyl-AMP ligase and continued by a multidomain non-ribosomal peptide synthetase/polyketide synthetase.High-resolution mass spectrometry and nuclear magnetic resonance spectroscopy measurements proved the production of puwainaphycin F/G congeners differing in FA chain length formed by either 3-amino-2-hydroxy-4-methyl dodecanoic acid (4-methyl-Ahdoa) or 3-amino-2-hydroxy-4-methyl tetradecanoic acid (4-methyl-Ahtea).Because only one puwainaphycin operon was recovered in the genome, we suggest that the fatty acyl-AMP ligase and one of the amino acid adenylation domains (Asn/Gln) show extended substrate specificity. View Article: PubMed Central - PubMed Affiliation: Institute of Microbiology AS CR, v.v.i., Department of Phototrophic Microorganisms - ALGATECH, Třeboň, Czech Republic; Biology Centre of AS CR, v.v.i., Institute of Hydrobiology, České Budějovice, Czech Republic; University of South Bohemia, Faculty of Science, Department of Botany, České Budějovice, Czech Republic. ABSTRACT A putative operon encoding the biosynthetic pathway for the cytotoxic cyanobacterial lipopeptides puwainphycins was identified in Cylindrospermum alatosporum. Bioinformatics analysis enabled sequential prediction of puwainaphycin biosynthesis; this process is initiated by the activation of a fatty acid residue via fatty acyl-AMP ligase and continued by a multidomain non-ribosomal peptide synthetase/polyketide synthetase. High-resolution mass spectrometry and nuclear magnetic resonance spectroscopy measurements proved the production of puwainaphycin F/G congeners differing in FA chain length formed by either 3-amino-2-hydroxy-4-methyl dodecanoic acid (4-methyl-Ahdoa) or 3-amino-2-hydroxy-4-methyl tetradecanoic acid (4-methyl-Ahtea). Because only one puwainaphycin operon was recovered in the genome, we suggest that the fatty acyl-AMP ligase and one of the amino acid adenylation domains (Asn/Gln) show extended substrate specificity. Our results provide the first insight into the biosynthesis of frequently occurring β-amino fatty acid lipopeptides in cyanobacteria, which may facilitate analytical assessment and development of monitoring tools for cytotoxic cyanobacterial lipopeptides. Show MeSH Major Bacterial Proteins/genetics/physiology Cyanobacteria/enzymology/genetics Ligases/genetics/physiology Polyketide Synthases/genetics/physiology Minor Biosynthetic Pathways Genes, Bacterial Lipopeptides/biosynthesis Molecular Sequence Annotation Molecular Sequence Data Multigene Family	© Copyright Policy Related In: Results - Collection License Show All Figures getmorefigures.php?uid=PMC4219810&req=5 pone-0111904-g001: Gene arrangement, functional annotation and domain structure of the puw gene cluster (56.7 kbp) from Cylindrospermum alatosporum CCALA 988. Mentions: After the final assembly of Illumina and Sanger sequencing data, the predicted puwainaphycin biosynthesis pathway was recovered in the middle of a ∼100-kbp long contig, with >20 kbp flanks at each side to ensure that the full pathway was sequenced. The putative biosynthetic gene cluster was 56,728 bp long and comprised 10 protein-coding ORFs. The ORFs were transcribed starting from a bi-directional promoter region, with orf2, puwA and orf1 transcribed in one direction and puwB-H in the opposite direction (Figure 1). The functional annotation of the individual enzymes and catalytic domains is summarized in Figure 1 and Table S3.

A hybrid non-ribosomal peptide/polyketide synthetase containing fatty-acyl ligase (FAAL) synthesizes the β-amino fatty acid lipopeptides puwainaphycins in the Cyanobacterium Cylindrospermum alatosporum.

Mareš J, Hájek J, Urajová P, Kopecký J, Hrouzek P - PLoS ONE (2014)

Bottom Line: Bioinformatics analysis enabled sequential prediction of puwainaphycin biosynthesis; this process is initiated by the activation of a fatty acid residue via fatty acyl-AMP ligase and continued by a multidomain non-ribosomal peptide synthetase/polyketide synthetase.High-resolution mass spectrometry and nuclear magnetic resonance spectroscopy measurements proved the production of puwainaphycin F/G congeners differing in FA chain length formed by either 3-amino-2-hydroxy-4-methyl dodecanoic acid (4-methyl-Ahdoa) or 3-amino-2-hydroxy-4-methyl tetradecanoic acid (4-methyl-Ahtea).Because only one puwainaphycin operon was recovered in the genome, we suggest that the fatty acyl-AMP ligase and one of the amino acid adenylation domains (Asn/Gln) show extended substrate specificity.

View Article: PubMed Central - PubMed

Affiliation: Institute of Microbiology AS CR, v.v.i., Department of Phototrophic Microorganisms - ALGATECH, Třeboň, Czech Republic; Biology Centre of AS CR, v.v.i., Institute of Hydrobiology, České Budějovice, Czech Republic; University of South Bohemia, Faculty of Science, Department of Botany, České Budějovice, Czech Republic.

ABSTRACT

A putative operon encoding the biosynthetic pathway for the cytotoxic cyanobacterial lipopeptides puwainphycins was identified in Cylindrospermum alatosporum. Bioinformatics analysis enabled sequential prediction of puwainaphycin biosynthesis; this process is initiated by the activation of a fatty acid residue via fatty acyl-AMP ligase and continued by a multidomain non-ribosomal peptide synthetase/polyketide synthetase. High-resolution mass spectrometry and nuclear magnetic resonance spectroscopy measurements proved the production of puwainaphycin F/G congeners differing in FA chain length formed by either 3-amino-2-hydroxy-4-methyl dodecanoic acid (4-methyl-Ahdoa) or 3-amino-2-hydroxy-4-methyl tetradecanoic acid (4-methyl-Ahtea). Because only one puwainaphycin operon was recovered in the genome, we suggest that the fatty acyl-AMP ligase and one of the amino acid adenylation domains (Asn/Gln) show extended substrate specificity. Our results provide the first insight into the biosynthesis of frequently occurring β-amino fatty acid lipopeptides in cyanobacteria, which may facilitate analytical assessment and development of monitoring tools for cytotoxic cyanobacterial lipopeptides.

Show MeSH

Gene arrangement, functional annotation and domain structure of the puw gene cluster (56.7 kbp) from Cylindrospermum alatosporum CCALA 988.

Related In: Results - Collection

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pone-0111904-g001: Gene arrangement, functional annotation and domain structure of the puw gene cluster (56.7 kbp) from Cylindrospermum alatosporum CCALA 988.

Mentions: After the final assembly of Illumina and Sanger sequencing data, the predicted puwainaphycin biosynthesis pathway was recovered in the middle of a ∼100-kbp long contig, with >20 kbp flanks at each side to ensure that the full pathway was sequenced. The putative biosynthetic gene cluster was 56,728 bp long and comprised 10 protein-coding ORFs. The ORFs were transcribed starting from a bi-directional promoter region, with orf2, puwA and orf1 transcribed in one direction and puwB-H in the opposite direction (Figure 1). The functional annotation of the individual enzymes and catalytic domains is summarized in Figure 1 and Table S3.