A photocrosslinking assay for reporting protein interactions in polyketide and fatty acid synthases

Mol Biosyst. 2011 Nov;7(11):3152-6. doi: 10.1039/c1mb05270e. Epub 2011 Sep 14.

Abstract

Understanding protein-protein interactions that occur between ACP and KS domains of polyketide synthases and fatty acid synthases is critical to improving the scope and efficiency of combinatorial biosynthesis efforts aimed at producing non-natural polyketides. Here, we report a facile strategy for rapidly reporting such ACP-KS interactions based on the incorporation of an amino acid with photocrosslinking functionality. Crucially, this photocrosslinking strategy can be applied to any polyketide or fatty acid synthase regardless of substrate specificity, and can be adapted to a high-throughput format for directed evolution studies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acyl Carrier Protein / chemistry
  • Acyl Carrier Protein / metabolism
  • Codon, Terminator
  • Cross-Linking Reagents / chemistry*
  • Cross-Linking Reagents / metabolism
  • Fatty Acid Synthases / chemistry*
  • Fatty Acid Synthases / metabolism
  • Kinetics
  • Mutagenesis, Site-Directed
  • Polyketide Synthases / chemistry*
  • Polyketide Synthases / metabolism
  • Polyketides / metabolism
  • Protein Conformation

Substances

  • Acyl Carrier Protein
  • Codon, Terminator
  • Cross-Linking Reagents
  • Polyketides
  • Polyketide Synthases
  • Fatty Acid Synthases