| Polyketides are a large class of structurally diverse and pharmacologically important natural products, which exhibit antibiotic, antitumor, anticancer, and other biological properties. In nature, polyketides are synthesized through repeated decarboxylative condensation from simple acyl-CoA precursors by polyketide synthases (PKSs). There are three types of PKSs. Type I PKSs are gigantic multifunctional enzymes that are organized into modules, each of which harbors a set of distinct, noniteratively acting activities responsible for the catalysis of one cycle of polyketide chain elongation. Type II PKSs are multienzyme complexes, in which the same set of enzymes are utilized in an iterative manner during the polyketide biosynthesis. In both cases, the growing polyketide chain is anchored on acyl carrier protein (ACP). Type III PKSs, also known as chalcone synthase-like PKSs, are structurally and mechanistically distinct from the first two types of PKSs. They are essentially condensing enzymes that lack ACP and act directly on acyl-CoA substrates.; Given the importance of polyketide natural products in human medicine, there is ongoing interest in producing unnatural polyketide products with novel or improved pharmacological properties. Whereas the structural complexity of polyketides makes their total organic synthesis impractical, biosynthesis through genetic engineering of PKSs is a promising route to produce novel polyketide analogues. Detailed structural insights into PKSs will greatly facilitate rational design of PKSs for successful novel polyketide production. As a step in this direction, the X-ray crystal structures of ketosynthase (KS)-acyltransferase (AT) didomains from module 3 and module 5 of 6-Deoxyerthronolide B synthase (DEBS), a prototypical type I PKS, were determined to 2.6 A and 2.7 A respectively. The overall fold of the two structures is homologous, with KS homodimer located in the center, flanked by the two AT monomers. In addition to the full length catalytic domains, the two structures also reveal three well-structured linker regions. Analysis of the substrate modeled active sites of KS3 and KS5 shows that a loop region might be in charge of the substrate specificity of KS domain. The distance between active sites of KS and AT is measured to be more than 70 A in both structures, thus domain reorganization is necessary for ACP to interact successfully with both KS and AT domains. The details of interaction between the two catalytic domains with ACP was investigated by docking simulation. ACP is proposed to dock within the deep cleft between KS and AT domains, with interactions span both the KS homodimer and AT domain.; The crystal structure of SCO1815, a NADPH-dependent ketoreductase (KR) was determined to 2.0 A. It is found to be functionally interact with the ACP from the initiation module of R1128 polyketide synthase, an extensively studied bimodular type II PKS. The structure shows that SCO1815 adopts a Rossmann fold and suggests that a conformational change occurs upon cofactor binding. We propose that a positively charged patch formed by three conserved residues is the ACP docking site. Our findings provide new engineering opportunities for incorporating unnatural primer units into novel polyketides. |
