Title: Structure and function of an iterative polyketide synthase thioesterase domain catalyzing Claisen cyclization in aflatoxin biosynthesis.
Authors: Korman, Tyler Paz; Crawford, Jason M; Labonte, Jason W; Newman, Adam G; Wong, Justin; Townsend, Craig A; Tsai, Shiou-Chuan
Published In Proc Natl Acad Sci U S A, (2010 Apr 06)
Abstract: Polyketide natural products possess diverse architectures and biological functions and share a subset of biosynthetic steps with fatty acid synthesis. The final transformation catalyzed by both polyketide synthases (PKSs) and fatty acid synthases is most often carried out by a thioesterase (TE). The synthetic versatility of TE domains in fungal nonreducing, iterative PKSs (NR-PKSs) has been shown to extend to Claisen cyclase (CLC) chemistry by catalyzing C-C ring closure reactions as opposed to thioester hydrolysis or O-C/N-C macrocyclization observed in previously reported TE structures. Catalysis of C-C bond formation as a product release mechanism dramatically expands the synthetic potential of PKSs, but how this activity was acquired has remained a mystery. We report the biochemical and structural analyses of the TE/CLC domain in polyketide synthase A, the multidomain PKS central to the biosynthesis of aflatoxin B(1), a potent environmental carcinogen. Mutagenesis experiments confirm the predicted identity of the catalytic triad and its role in catalyzing the final Claisen-type cyclization to the aflatoxin precursor, norsolorinic acid anthrone. The 1.7 A crystal structure displays an alpha/beta-hydrolase fold in the catalytic closed form with a distinct hydrophobic substrate-binding chamber. We propose that a key rotation of the substrate side chain coupled to a protein conformational change from the open to closed form spatially governs substrate positioning and C-C cyclization. The biochemical studies, the 1.7 A crystal structure of the TE/CLC domain, and intermediate modeling afford the first mechanistic insights into this widely distributed C-C bond-forming class of TEs.
PubMed ID: 20332208
MeSH Terms: Aflatoxins/biosynthesis*; Biocatalysis; Crystallography, X-Ray; Cyclization; Fungal Proteins/chemistry*; Fungal Proteins/metabolism*; Hydrophobic and Hydrophilic Interactions; Mutation; Polyketide Synthases/chemistry*; Polyketide Synthases/genetics; Polyketide Synthases/metabolism*; Protein Folding; Protein Structure, Tertiary; Thiolester Hydrolases/chemistry*; Thiolester Hydrolases/metabolism*