Structural Analysis of Actinorhodin Polyketide Ketoreductase: Cofactor Binding and Substrate Specificity

Abstract

Aromatic polyketides are a class of natural products that include many pharmaceutically important aromatic compounds. Understanding the structure and function of PKS will provide clues to the molecular basis of polyketide biosynthesis. Polyketide chain reduction by ketoreductase (KR) provides regio- and stereochemical diversity. Cocrystal structures of actinorhodin polyketide ketoreductase (act KR) were solved to 2.3 Å with either NADP+ or NADPH bound. The monomer fold is a conserved Rossmann fold. Small structural differences between act KR and fatty acid KRs fine-tune the tetramer interface and substrate binding pocket. Comparison of the NADP+- and NADPH-bound structures indicate that the α6-α7 loop region is highly flexible. The intricate proton-relay network in the active site leads to the proposed catalytic mechanism involving four waters, NADPH, and residues N114-S144-Y157-K161. Acyl carrier protein and substrate docking models shed light on the molecular basis of KR regio- and stereoselectivity. Sequence comparison indicates that the above features are highly conserved among aromatic polyketide KRs. The structures of act KR provide an important step toward understanding aromatic PKS and will enhance our ability to design novel aromatic polyketide natural products with different reduction patterns.