Exploration of polyyne biosynthetic gene cluster diversity in bacteria leads to the discovery of thePseudomonaspolyyne protegencin

Abstract

Natural products that possess alkyne or polyyne moieties have been isolated from a variety of biological sources. In bacteria their biosynthesis has been defined, however, the distribution of polyyne biosynthetic gene clusters (BGCs), and their evolutionary relationship to alkyne biosynthesis, have not been addressed. We explored the distribution of alkyne biosynthesis gene cassettes throughout bacteria, revealing evidence of multiple horizontal gene transfer events. Investigating the evolutionary connection between alkyne and polyyne biosynthesis identified a monophyletic clade possessing a conserved seven-gene cassette for polyyne biosynthesis. Mapping the diversity of these conserved genes revealed a phylogenetic clade representing a polyyne BGC in Pseudomonas, pgn, and subsequent pathway mutagenesis and analytical chemistry characterised the associated metabolite, protegencin. In addition to unifying and expanding our knowledge of polyyne diversity, our results show that alkyne and polyyne biosynthetic gene clusters are promiscuous within bacteria. Systematic mapping of conserved biosynthetic genes across bacterial genomic diversity has proven to be a successful method for discovering natural products.

A polyyne toxin produced by an antagonistic bacterium blinds and lyses a Chlamydomonad alga

“…P. protegens produces a variety of secondary metabolites controlled by the GacS-GacA regulatory system ( 16 , 34 , 40 ), including the pgn cluster. The production of a polyyne was predicted in silico ( 31 ) and recently verified in Pseudomonas ( 33 , SI Appendix , SI Note ). Polyynes like protegencin are notoriously unstable compounds.…”

“…In bacteria, polyyne biosynthesis is encoded in gene clusters bearing genes for desaturases, a fatty acyl-AMP ligase, an acyl carrier protein, and a thioesterase ( 31 ). Mining the genome ( 32 ) of P. protegens revealed a tentative polyyne biosynthesis gene cluster (pgn) for a compound named “protegencin” ( 33 ). According to homology searches, pgnE , pgnF , and pgnH encode desaturases and a thioesterase; pgnD encodes a fatty acyl-AMP ligase, and pgnG encodes an acyl carrier protein ( Fig.…”

“…In a previous genome mining analysis, orthologous pgn gene clusters were also found in Collimonas and Mycobacterium as well as in different Burkholderia species ( 31 ). Recently, the distribution of the pgn cluster was found in over 50 different Pseudomonas genomes ( 33 ). Currently, it is not known whether all these bacteria can also exert algicidal activities using polyynes; the interactions of these bacteria have also only been studied with organisms other than photosynthetic protists.…”