PRODUCTION OF NEW WAP-8294A CYCLODEPSIPEPTIDES BY THE BIOLOGICAL CONTROL AGENT LYSOBACTER ENZYMOGENES OH11

● Lysobacter enzymogenes mutants were generated for WAP-8294A biosynthesis. ● Essential and non-essential accessory genes for WAP-8294A biosynthesis were determined. ● Six new WAP-8294A analogs were identified using UHPLC-HR-MS/MS. ● Three deoxy analogs were detected supporting the function of ORF4 in asparagine hydroxylation. Naturally-occurring environmental microorganisms may provide ‘green’ and effective biocontrol tools for disease management in agricultural crops. Due to the constant threat of resistant pathogens there is a pressing and continual need to search for new biocontrol tools. This study investigated the production of new analogs of WAP-8294A compounds by the biocontrol agent Lysobacter enzymogenes OH11 through biosynthetic engineering. WAP-8294As are a family of natural cyclic lipodepsipeptides with potent activity against Gram-positive bacteria. A series of genetic manipulations was therefore conducted on the accessory genes in the WAP biosynthetic gene cluster. The resulting strains containing a single-point mutation in ORF4, which was predicted to encode a 2-ketoglutarate dependent dioxygenase, produced deoxy-WAP-8294As. This result provides evidence for the function of ORF4 in catalyzing β-hydroxylation of the D-asparagine residue in WAP-8294As. In addition, six new analogs of WAP-8294As were identified by UHPLC-HR-MS/MS. This is the first attempt to produce new WAP-8294As in Lysobacter and shows that the spectrum of the biocontrol compounds may be expanded through the manipulation of biosynthetic genes.