REACTION-MECHANISMS OF HOMODIMERIC PLANT POLYKETIDE SYNTHASES (STILBENE AND CHALCONE SYNTHASE) - A SINGLE ACTIVE-SITE FOR THE CONDENSING REACTION IS SUFFICIENT FOR SYNTHESIS OF STILBENES, CHALCONES, AND 6'-DEOXYCHALCONES

Stilbene (STS) and chalcone (CHS) synthases are homodimeric, related p lant-specific polyketide synthases. Both perform a sequential condensa tion of three acetate units to a starter residue to form a tetraketide intermediate that is folded to the ring systems specific to the diffe rent products. Protein cross-linking and site-directed mutagenesis ide ntified a subunit contact site in position 158, close to the active si te (Cys(169)). This suggested that the active site pockets may be neig hboring, possibly alternating in the condensing reactions rather than acting independently. This was investigated by coexpression of active site mutants with differently mutated, inactive proteins. With both ST S and CHS, the heterodimers synthesized the end products, indicating t hat each subunit performed all three condensations. In co-action with a monomeric reductase, CHS also synthesizes 6'-deoxychalcone, but with the chalcone as second product when using plant preparations. The two enzymes expressed as single species in Escherichia coli synthesized b oth products, and both were also obtained with a CHS heterodimer conta ining a single active site. The results showed that 6'-deoxychalcone s ynthesis required no other plant factor and that the formation of two products may be an intrinsic property of the interaction between dimer ic CHS and monomeric reductase.