CHARACTERIZATION OF THE GENES-CONTROLLING THE BIOSYNTHESIS OF THE POLYKETIDE PHYTOTOXIN CORONATINE INCLUDING CONJUGATION BETWEEN CORONAFACIC AND CORONAMIC ACID

Pseudomonas syringae pv. glycinea PG4180 produces a chlorosis-inducing phytotoxin, coronatine (COR), which consists of a polyketide componen t, coronafacic acid (CFA), which is coupled via amide bond formation t o coronamic acid (CMA), an ethylcyelopropyl amino acid (aa) derived fr om isoleucine. P. syringae pv. syringae strains PS51 and PS61, which d o not synthesize coronafacoyl compounds (conjugates between CFA and aa ), acquired the ability to produce CFA and COR when transformed with p 4180A, a 90-kb indigenous plasmid in PG4180. Tn5 mutagenesis indicated that the COR biosynthetic genes in PG4180 are clustered within a 30-k b region on p4180A. The phenotype of selected COR-defective mutants wa s determined by supplying them with CFA and CMA and by complementation studies with cloned DNA from the COR biosynthetic cluster. Using this approach, the regions encoding CFA and CMA synthesis and coupling act ivity were localized to the 24-, 12.5- and 2.3-kb regions of the clust er, respectively. Mutants in a 6-kb region required the addition of bo th CFA and CMA for COR synthesis, which may indicate a regulatory role for this part of the cluster. PS51 and PS61 transconjugants containin g cloned DNA from the coupling region produced COR when supplied with CFA and CMA, indicating that coupling activity was cloned and expresse d in bacteria lacking the COR biosynthetic cluster.