The genomisotopic approach: a systematic method to isolate products of orphan biosynthetic gene clusters.
With the increasing number of genomes sequenced and available in the public domain, a large number of orphan gene clusters, for which the encoded natural product is unknown, have been identified. These orphan gene clusters represent a tremendous source of novel and possibly bioactive compounds. Here, we describe a "genomisotopic approach," which employs a combination of genomic sequence analysis and isotope-guided fractionation to identify unknown compounds synthesized from orphan gene clusters containing nonribosomal peptide synthetases. Analysis of the Pseudomonas fluorescens Pf-5 genome led to the identification of an orphan gene cluster predicted to code for the biosynthesis of a lipopeptide natural product. Application of the genomisotopic approach to isolate the product of this gene cluster resulted in the discovery of orfamide A, founder of a group of bioactive cyclic lipopeptides.
Gross
Harald
H
Center for Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography and Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
Stockwell
Virginia O
VO
Henkels
Marcella D
MD
Nowak-Thompson
Brian
B
Loper
Joyce E
JE
Gerwick
William H
WH
D016680:Genome, Bacterial / Q000235:genetics*; D023281:Genomics / Q000379:methods*; D007554:Isotopes; D008074:Lipoproteins / Q000096:biosynthesis / Q000302:isolation & purification; D008722:Methods; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D010453:Peptide Synthases / Q000235:genetics; D010456:Peptides, Cyclic / Q000096:biosynthesis / Q000302:isolation & purification; D011551:Pseudomonas fluorescens / Q000235:genetics; D017421:Sequence Analysis
D007554:Isotopes; D008074:Lipoproteins; D010456:Peptides, Cyclic; D010453:Peptide Synthases