Structures and Biosynthesis of Caryoynencins, Unstable Bacterial Polyynes from Pseudomonas protegens Recombinant Expressing the cayG Gene

Bacteria in certain genera can produce “bacterial polyynes” that contain a conjugated CC bond starting from a terminal alkyne. Protegenin A is a derivative of octadecanoic acid that contains an ene–tetrayne moiety. It was discovered in Pseudomonas protegens Cab57 and exhibits strong antioomycete and moderate antifungal activity. By introducing cayG, a cytochrome P450 gene from Burkholderia caryophylli, into P. protegens Cab57, protegenin A was converted into more complex polyynes, caryoynencins A–E. A purification method that minimized the degradation and isomerization of caryoynencins was established. For the first time, as far as we know, the ¹H and ¹³C{¹H} NMR signals of caryoynencins were completely assigned by analyzing the NMR data of the isolated compounds and protegenin A enriched with [1-¹³C]- or [2-¹³C]-acetate. Through the structural analysis of caryoynencins D/E and bioconversion experiments, we observed that CayG constructs the allyl alcohol moiety of caryoynencins A–C through sequential hydroxylation, dehydration, and hydroxylation. The recombinant strain exhibited a stronger antioomycete activity compared to the wild-type strain. This paper proposes a stable purification and structural determination method for various bacterial polyynes, and P. protegens Cab57 holds promise as an engineering host for the production of biologically active polyynes.