Structure and Function of an Iterative Polyketide Synthase Thioesterase Domain Catalyzing Claisen Cyclization in Aflatoxin Biosynthesis

Proc Natl Acad Sci U S A. 2010 Apr 6;107(14):6246-51. doi: 10.1073/pnas.0913531107. Epub 2010 Mar 23.


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

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*


  • Aflatoxins
  • Fungal Proteins
  • Polyketide Synthases
  • Thiolester Hydrolases

Associated data

  • PDB/3D4H