Enzymatic formation of an aromatic dodecaketide by engineered plant polyketide synthase

Octaketide synthase (OKS) from Aloe arborescens is a plant-specific type III polyketide synthase (PKS) that catalyzes iterative condensations of eight molecules of malonyl-CoA to produce the C(16) aromatic octaketides SEK4 and SEK4b. On the basis of the crystal structures of OKS, the F66L/N222G double mutant was constructed and shown to produce an unnatural dodecaketide TW95a by sequential condensations of 12 molecules of malonyl-CoA. The C(24) naphthophenone TW95a is a product of the minimal type II PKS (whiE from Streptomyces coelicolor), and is structurally related to the C(20) decaketide benzophenone SEK15, the product of the OKS N222G point mutant. The C(24) dodecaketide naphthophenone TW95a is the first and the longest polyketide scaffold generated by a structurally simple type III PKS. A homology model predicted that the active-site cavity volume of the F66L/N222G mutant is increased to 748Å(3), from 652Å(3) of the wild-type OKS. The structure-based engineering thus greatly expanded the catalytic repertoire of the simple type III PKS to further produce larger and more complex polyketide molecules.