Updated project metadata. The molecular chaperone heat shock protein 90 (HSP90) works in concert with its co-chaperones to stabilize its client proteins, which include several drivers of oncogenesis and malignant progression. Pharmacologic inhibitors of HSP90 are proposed to trigger widespread remodeling of cellular protein complexes, including dissociation of co-chaperones from HSP90, disruption of client protein signaling networks, and recruitment of the protein ubiquitination and degradation machinery. However, proteomic studies to date have focused on inhibitor-induced changes in total protein levels, often overlooking protein complex alterations. Here, we use size-exclusion chromatography in combination with mass spectrometry (SEC-MS) to characterize the changes in native protein complexes following treatment with the HSP90 inhibitor 17-AAG in the human HT29 colon adenocarcinoma cell line.