1. Structure and function of an iterative polyketide synthase thioesterase domain catalyzing Claisen cyclization in aflatoxin biosynthesis
- Author
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Korman, Tyler Paz, Crawford, Jason M., Labonte, Jason W., Newman, Adam G., Wong, Justin, Townsend, Craig A., and Tsai, Shiou-Chuan
- Subjects
Aflatoxins -- Health aspects ,Aflatoxins -- Research ,Esterases -- Physiological aspects ,Esterases -- Structure ,Esterases -- Research ,Fatty acids -- Synthesis ,Fatty acids -- Physiological aspects ,Fatty acids -- Research ,Science and technology - 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 B1, 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 [Angstrom] crystal structure displays an a/[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 [Angstrom] 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. biosynthesis | fungal metabolism | mutagenesis doi/10.1073/pnas.0913531107
- Published
- 2010