Photoreactive Stapled BH3 Peptides to Dissect the BCL-2 Family Interactome

Department of Pediatric Oncology, Dana-Farber Cancer Institute and Children's Hospital Boston, Harvard Medical School, Boston, MA 02115, USA.
Chemistry & biology (Impact Factor: 6.65). 12/2010; 17(12):1325-33. DOI: 10.1016/j.chembiol.2010.09.015
Source: PubMed


Defining protein interactions forms the basis for discovery of biological pathways, disease mechanisms, and opportunities for therapeutic intervention. To harness the robust binding affinity and selectivity of structured peptides for interactome discovery, we engineered photoreactive stapled BH3 peptide helices that covalently capture their physiologic BCL-2 family targets. The crosslinking α helices covalently trap both static and dynamic protein interactors, and enable rapid identification of interaction sites, providing a critical link between interactome discovery and targeted drug design.

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    • "The eluates were electrophoresed using 4%–12% gradient Bis-Tris gels (Invitrogen) and then stained for protein content by Coomassie blue. The crosslinked bands were excised and prepared for mass spectrometry analysis as previously described (Braun et al., 2010). Samples were subjected to liquid chromatography-tandem mass spectrometry (LC-MS/MS) in an LTQ Orbitrap XL hybrid mass spectrometer (Thermo Fisher Scientific). "
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