Knowledge-based design of bimodular and trimodular polyketide synthases based on domain and module swaps: a route to simple statin analogues

Background: Polyketides are structurally diverse natural products that have a range of medically useful activities, Nonaromatic bacterial polyketides are synthesised on modular polyketide synthase (PKS) multienzymes, in which each cycle of chain extension requires a different 'module' of enzymatic a ctivities. Attempts to design and construct modular PKSs that synthesise sp ecified novel polyketides provide a particularly stringent test of our unde rstanding of PKS structure and function. Results: We have constructed bimodular and trimodular PKSs based on DEBS1-T E, a derivative of the erythromycin PKS that contains only modules 1 and 2 and a thioesterase (TE), by substituting multiple domains with appropriate counterparts derived from the rapamycin PKS. Hybrid PKSs were obtained that synthesised the predicted target triketide lactones, which are simple anal ogues of cholesterol-lowering statins, In constructing intermodular fusions , whether between modules in the same or in different proteins, it was foun d advantageous to preserve intact the acyl carrier protein-ketosynthase (AC P-KS) didomain that spans the junction between successive modules. Conclusions: Relatively simple considerations govern the construction of fu nctional hybrid PKSs, Fusion sites should be chosen either in the surface-a ccessible linker regions between enzymatic domains, as previously revealed, or just inside the conserved margins of domains. The interaction of an ACP domain with the adjacent KS domain, whether on the same polyketide or not, is of particular importance, both through conservation of appropriate prot ein-protein interactions, and through optimising molecular recognition of t he altered polyketide chain in the key transfer of the acyl chain from the ACP of one module to the KS of the downstream module.