Reaction mechanisms of homodimeric plant polyketide synthase (stilbenes and chalcone synthase). A single active site for the condensing reaction is sufficient for synthesis of stilbenes, chalcones, and 6'-deoxychalcones

Abstract

Stilbene (STS) and chalcone (CHS) synthases are homodimeric, related plant-specific polyketide synthases. Both perform a sequential condensation of three acetate units to a starter residue to form a tetraketide intermediate that is folded to the ring systems specific to the different products. Protein cross-linking and site-directed mutagenesis identified a subunit contact site in position 158, close to the active site (Cys169). This suggested that the active site pockets may be neighboring, 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 STS and CHS, the heterodimers synthesized the end products, indicating that 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 both products, and both were also obtained with a CHS heterodimer containing a single active site. The results showed that 6'-deoxychalcone synthesis required no other plant factor and that the formation of two products may be an intrinsic property of the interaction between dimeric CHS and monomeric reductase.