title: A highly selective Hsp90 affinity chromatography resin with a cleavable linker.
abstract: Over 200 proteins have been identified that interact with the protein chaperone Hsp90, a recognized therapeutic target thought to participate in non-oncogene addiction in a variety of human cancers. However, defining Hsp90 clients is challenging because interactions between Hsp90 and its physiologically relevant targets involve low affinity binding and are thought to be transient. Using a chemo-proteomic strategy, we have developed a novel orthogonally cleavable Hsp90 affinity resin that allows purification of the native protein and is quite selective for Hsp90 over its immediate family members, GRP94 and TRAP 1. We show that the resin can be used under low stringency conditions for the rapid, unambiguous capture of native Hsp90 in complex with a native client. We also show that the choice of linker used to tether the ligand to the insoluble support can have a dramatic effect on the selectivity of the affinity media.
authors.affiliation: Department of Pharmacology and Cancer Biology, C119 LSRC, Research Drive, Duke University, Durham, NC 27710, United States.
authors.forename: Philip F
authors.initials: PF
authors.lastname: Hughes
authors.forename: Jared J
authors.initials: JJ
authors.lastname: Barrott
authors.forename: David A
authors.initials: DA
authors.lastname: Carlson
authors.forename: David R
authors.initials: DR
authors.lastname: Loiselle
authors.forename: Brittany L
authors.initials: BL
authors.lastname: Speer
authors.forename: Khaldon
authors.initials: K
authors.lastname: Bodoor
authors.forename: Lauretta A
authors.initials: LA
authors.lastname: Rund
authors.forename: Timothy A J
authors.initials: TA
authors.lastname: Haystead
mesh_terms: D000818:Animals; D002846:Chromatography, Affinity / Q000295:instrumentation*; D004591:Electrophoresis, Polyacrylamide Gel; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D011485:Protein Binding; D040901:Proteomics; D012117:Resins, Synthetic / Q000737:chemistry* / Q000378:metabolism*; D012680:Sensitivity and Specificity; D013552:Swine
chemical_list: D018841:HSP90 Heat-Shock Proteins; D012117:Resins, Synthetic