title: A genome-wide analysis of nonribosomal peptide synthetase gene clusters and their peptides in a Planktothrix rubescens strain.
abstract: BACKGROUND
  Cyanobacteria often produce several different oligopeptides, with unknown biological functions, by nonribosomal peptide synthetases (NRPS). Although some cyanobacterial NRPS gene cluster types are well described, the entire NRPS genomic content within a single cyanobacterial strain has never been investigated. Here we have combined a genome-wide analysis using massive parallel pyrosequencing ("454") and mass spectrometry screening of oligopeptides produced in the strain Planktothrix rubescens NIVA CYA 98 in order to identify all putative gene clusters for oligopeptides.
  
  
  RESULTS
  Thirteen types of oligopeptides were uncovered by mass spectrometry (MS) analyses. Microcystin, cyanopeptolin and aeruginosin synthetases, highly similar to already characterized NRPS, were present in the genome. Two novel NRPS gene clusters were associated with production of anabaenopeptins and microginins, respectively. Sequence-depth of the genome and real-time PCR data revealed three copies of the microginin gene cluster. Since NRPS gene cluster candidates for microviridin and oscillatorin synthesis could not be found, putative (gene encoded) precursor peptide sequences to microviridin and oscillatorin were found in the genes mdnA and oscA, respectively. The genes flanking the microviridin and oscillatorin precursor genes encode putative modifying enzymes of the precursor oligopeptides. We therefore propose ribosomal pathways involving modifications and cyclisation for microviridin and oscillatorin. The microviridin, anabaenopeptin and cyanopeptolin gene clusters are situated in close proximity to each other, constituting an oligopeptide island.
  
  
  CONCLUSIONS
  Altogether seven nonribosomal peptide synthetase (NRPS) gene clusters and two gene clusters putatively encoding ribosomal oligopeptide biosynthetic pathways were revealed. Our results demonstrate that whole genome shotgun sequencing combined with MS-directed determination of oligopeptides successfully can identify NRPS gene clusters and the corresponding oligopeptides. The analyses suggest independent evolution of all NRPS gene clusters as functional units. Our data indicate that the Planktothrix genome displays evolution of dual pathways (NRPS and ribosomal) for production of oligopeptides in order to maximize the diversity of oligopeptides with similar but functional discrete bioactivities.
authors.affiliation: University of Oslo, Department of Biology, Centre for Ecological and Evolutionary Synthesis (CEES), 0316 Oslo, Norway. t.b.rounge@bio.uio.no
authors.forename: Trine B
authors.initials: TB
authors.lastname: Rounge
authors.forename: Thomas
authors.initials: T
authors.lastname: Rohrlack
authors.forename: Alexander J
authors.initials: AJ
authors.lastname: Nederbragt
authors.forename: Tom
authors.initials: T
authors.lastname: Kristensen
authors.forename: Kjetill S
authors.initials: KS
authors.lastname: Jakobsen
mesh_terms: D019295:Computational Biology; D000458:Cyanobacteria / Q000235:genetics* / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics; D019143:Evolution, Molecular; D005798:Genes, Bacterial; D016680:Genome, Bacterial; D055106:Genome-Wide Association Study; D005810:Multigene Family; D009842:Oligopeptides / Q000096:biosynthesis; D010453:Peptide Synthases / Q000235:genetics*; D010802:Phylogeny; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D004269:DNA, Bacterial; D009842:Oligopeptides; D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase
references.citation: Microbiology (Reading). 2007 May;153(Pt 5):1382-1393
references.pmid: 17464052
references.citation: Appl Environ Microbiol. 2004 Feb;70(2):686-92
references.pmid: 14766543
references.citation: Proc Natl Acad Sci U S A. 2007 Aug 28;104(35):14092-7
references.pmid: 17715057
references.citation: Nucleic Acids Res. 2009 Jan;37(Database issue):D211-5
references.pmid: 18940856
references.citation: Mol Microbiol. 2000 Jul;37(1):156-67
references.pmid: 10931313
references.citation: Appl Environ Microbiol. 2008 Mar;74(6):1791-7
references.pmid: 18245249
references.citation: Appl Environ Microbiol. 2001 Nov;67(11):5069-76
references.pmid: 11679328
references.citation: J Nat Prod. 2007 Jul;70(7):1084-8
references.pmid: 17602586
references.citation: Chem Rev. 2005 Feb;105(2):715-38
references.pmid: 15700962
references.citation: Bioinformatics. 2003 Aug 12;19(12):1572-4
references.pmid: 12912839
references.citation: Chem Biol. 2000 Oct;7(10):753-64
references.pmid: 11033079
references.citation: Genetics. 1999 Dec;153(4):1525-33
references.pmid: 10581263
references.citation: FEMS Microbiol Rev. 2006 Jul;30(4):530-63
references.pmid: 16774586
references.citation: FEMS Microbiol Lett. 2001 Mar 15;196(2):207-14
references.pmid: 11267781
references.citation: J Bacteriol. 2003 May;185(9):2774-85
references.pmid: 12700256
references.citation: Chem Biol. 1999 Aug;6(8):493-505
references.pmid: 10421756
references.citation: Nucleic Acids Res. 2001 Jan 1;29(1):173-4
references.pmid: 11125082
references.citation: Brief Bioinform. 2004 Jun;5(2):150-63
references.pmid: 15260895
references.citation: J Bacteriol. 2003 Jan;185(2):564-72
references.pmid: 12511503
references.citation: Proc Natl Acad Sci U S A. 2005 May 17;102(20):7315-20
references.pmid: 15883371
references.citation: BMC Microbiol. 2008 Aug 26;8:141
references.pmid: 18727817
references.citation: Appl Environ Microbiol. 2004 Nov;70(11):6370-8
references.pmid: 15528494
references.citation: Mol Ecol. 2006 Oct;15(12):3849-61
references.pmid: 17032280
references.citation: Angew Chem Int Ed Engl. 2008;47(40):7756-9
references.pmid: 18683268
references.citation: Bioinformatics. 2005 May 1;21(9):2104-5
references.pmid: 15647292
references.citation: Proc Natl Acad Sci U S A. 2001 Oct 9;98(21):12215-20
references.pmid: 11572948
references.citation: J Nat Prod. 1996 Jan;59(1):90-2
references.pmid: 8984160
references.citation: Chem Biol. 2000 Mar;7(3):211-24
references.pmid: 10712928
references.citation: Bioorg Med Chem Lett. 1999 May 3;9(9):1243-6
references.pmid: 10340607
references.citation: Bioinformatics. 1998;14(1):68-73
references.pmid: 9520503
references.citation: Genetics. 2006 Apr;172(4):2665-81
references.pmid: 16489234
references.citation: Microbiology (Reading). 2004 Jun;150(Pt 6):1785-1796
references.pmid: 15184565
references.citation: Chem Rev. 1997 Nov 10;97(7):2651-2674
references.pmid: 11851476
references.citation: Appl Environ Microbiol. 2007 Nov;73(22):7322-30
references.pmid: 17921284
references.citation: Microbiology (Reading). 2005 May;151(Pt 5):1525-1533
references.pmid: 15870462
references.citation: Chem Biol. 2007 May;14(5):565-576
references.pmid: 17524987
references.citation: J Chem Ecol. 2003 Aug;29(8):1757-70
references.pmid: 12956505
references.citation: FEMS Microbiol Lett. 2005 Nov 1;252(1):143-51
references.pmid: 16182472
references.citation: J Mol Biol. 1990 Oct 5;215(3):403-10
references.pmid: 2231712
references.citation: Nucleic Acids Res. 2007 Jan;35(Database issue):D169-72
references.pmid: 17090583
references.citation: J Mol Evol. 2004 Jun;58(6):633-41
references.pmid: 15461420
references.citation: Appl Environ Microbiol. 2004 Aug;70(8):5047-50
references.pmid: 15294849