Drug Affinity Responsive Target Stability (DARTS) to Resolve Protein–Small Molecule Interaction in Arabidopsis

Abstract

Target identification remains a challenging step in plant chemical genomics approaches. Drug affinity responsive target stability (DARTS) represents a straightforward technique to identify small molecules’ protein targets and assist in the characterization of interactions between small molecules and putative targets identified by other methods. When a small molecule interacts with a protein, it has the potential to stabilize the protein's structure, resulting in a reduced susceptibility to protease action. During the DARTS procedure, protein extracts are treated with proteolytic enzymes, and only proteins that bind to the small molecule are protected from proteolysis. DARTS represents a protocol independent of the molecule's mechanism of action or chemical structure. Another advantage of DARTS is that it does not require additional modifications or tagging of the small molecule. The protocols outlined in this article describe in detail the DARTS technique applied to plant proteins and propose several detection procedures according to protein abundance. © 2017 by John Wiley & Sons, Inc.

Discovery of Pyruvate Kinase as a Novel Target of New Fungicide Candidate 3-(4-Methyl-1,2,3-thiadiazolyl)-6-trichloromethyl-[1,2,4]-triazolo-[3,4-b][1,3,4]-thiadizole

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“…The DARTS assay was performed as previously described (35,37). See SI Appendix, Supplemental Materials and Methods.…”

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