title: Molecular characterization of the safracin biosynthetic pathway from Pseudomonas fluorescens A2-2: designing new cytotoxic compounds.
abstract: Safracin is an antibiotic with anti-tumour activity produced by Pseudomonas fluorescens A2-2. The entire safracin synthetic gene cluster spanning 17.5 kb has been identified, cloned and sequenced. The safracin cluster comprises 10 open reading frames (ORFs) encoding proteins for three non-ribosomal peptide synthetases (NRPS), three safracin precursor biosynthetic enzymes, two safracin tailoring enzymes, a safracin resistance protein and a small hypothetical protein of unknown function. These genes are organized in two divergent operons of eight and two genes respectively. This pathway exhibits unusual features when compared with other NRPS systems. We have demonstrated by heterologous expression of the cluster that it is able to direct the synthesis of safracin in other strains. Cross-feeding experiments have confirmed that 3-hydroxy-5-methyl-O-methyltyrosine is the precursor of two amino acids of the molecule. Genetic analyses have allowed us to demonstrate that the bicistronic operon encodes the hydroxylation and N-methylation activities of the pathway. The cloning and expression of the safracin cluster has settled the basis for the in vivo and in vitro production of a wide variety of compounds, such as the promising ecteinascidins anti-cancer compounds.
authors.affiliation: Drug Discovery Department, PharmaMar, S.A. E-28770-Colmenar Viejo, Madrid, Spain. avelaco@pharmamar.com
authors.forename: Ana
authors.initials: A
authors.lastname: Velasco
authors.forename: Paloma
authors.initials: P
authors.lastname: Acebo
authors.forename: Alicia
authors.initials: A
authors.lastname: Gomez
authors.forename: Carmen
authors.initials: C
authors.lastname: Schleissner
authors.forename: Pilar
authors.initials: P
authors.lastname: Rodríguez
authors.forename: Tomas
authors.initials: T
authors.lastname: Aparicio
authors.forename: Susana
authors.initials: S
authors.lastname: Conde
authors.forename: Rosario
authors.initials: R
authors.lastname: Muñoz
authors.forename: Fernando
authors.initials: F
authors.lastname: de la Calle
authors.forename: Jose Luis
authors.initials: JL
authors.lastname: Garcia
authors.forename: Jose M
authors.initials: JM
authors.lastname: Sánchez-Puelles
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D024881:Drug Resistance, Bacterial; D007546:Isoquinolines / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D010453:Peptide Synthases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011551:Pseudomonas fluorescens / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D017422:Sequence Analysis, DNA
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D007546:Isoquinolines; D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase