Proteomic discovery of chemical probes that perturb protein complexes in human cells

  • Mol Cell. 2023 May 18;83(10):1725-1742.e12. doi: 10.1016/j.molcel.2023.03.026.
Michael R Lazear  1 ,  Jarrett R Remsberg  2 ,  Martin G Jaeger  1 ,  Katherine Rothamel  3 ,  Hsuan-Lin Her  3 ,  Kristen E DeMeester  1 ,  Evert Njomen  1 ,  Simon J Hogg  4 ,  Jahan Rahman  4 ,  Landon R Whitby  5 ,  Sang Joon Won  1 ,  Michael A Schafroth  1 ,  Daisuke Ogasawara  1 ,  Minoru Yokoyama  1 ,  Garrett L Lindsey  1 ,  Haoxin Li  1 ,  Jason Germain  1 ,  Sabrina Barbas  1 ,  Joan Vaughan  6 ,  Thomas W Hanigan  1 ,  Vincent F Vartabedian  7 ,  Christopher J Reinhardt  1 ,  Melissa M Dix  1 ,  Seong Joo Koo  8 ,  Inha Heo  8 ,  John R Teijaro  7 ,  Gabriel M Simon  5 ,  Brahma Ghosh  9 ,  Omar Abdel-Wahab  4 ,  Kay Ahn  10 ,  Alan Saghatelian  6 ,  Bruno Melillo  11 ,  Stuart L Schreiber  12 ,  Gene W Yeo  3 ,  Benjamin F Cravatt  13
Affiliations
  • 1. Department of Chemistry, Scripps Research, La Jolla, CA 92037, USA.
  • 2. Department of Chemistry, Scripps Research, La Jolla, CA 92037, USA. Electronic address: [email protected].
  • 3. Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA, USA.
  • 4. Human Oncology and Pathogenesis Program, Memorial Sloan Kettering Cancer Center, New York, NY 10021, USA.
  • 5. Vividion Therapeutics, 5820 Nancy Ridge Drive, San Diego, CA 92121, USA.
  • 6. Clayton Foundation Laboratories for Peptide Biology, The Salk Institute for Biological Studies, La Jolla, CA, USA.
  • 7. Department of Immunology and Microbiology, Scripps Research, La Jolla, CA 92037, USA.
  • 8. Molecular and Cellular Pharmacology, Discovery Technologies and Molecular Pharmacology, Janssen Research and Development, Turnhoutseweg 30, 2340 Beerse, Belgium.
  • 9. Discovery Chemistry, Janssen Research & Development, Spring House, PA 19477, USA.
  • 10. Molecular and Cellular Pharmacology, Discovery Technologies and Molecular Pharmacology, Janssen Research and Development, Spring House, PA 19477, USA.
  • 11. Department of Chemistry, Scripps Research, La Jolla, CA 92037, USA; Chemical Biology and Therapeutics Science Program, Broad Institute, Cambridge, MA 02142, USA.
  • 12. Chemical Biology and Therapeutics Science Program, Broad Institute, Cambridge, MA 02142, USA; Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138, USA.
  • 13. Department of Chemistry, Scripps Research, La Jolla, CA 92037, USA. Electronic address: [email protected].
PMID: 37084731 DOI: 10.1016/j.molcel.2023.03.026
Abstract

Most human proteins lack chemical probes, and several large-scale and generalizable small-molecule binding assays have been introduced to address this problem. How compounds discovered in such "binding-first" assays affect protein function, nonetheless, often remains unclear. Here, we describe a "function-first" proteomic strategy that uses size exclusion chromatography (SEC) to assess the global impact of electrophilic compounds on protein complexes in human cells. Integrating the SEC data with cysteine-directed activity-based protein profiling identifies changes in protein-protein interactions that are caused by site-specific liganding events, including the stereoselective engagement of cysteines in PSME1 and SF3B1 that disrupt the PA28 Proteasome regulatory complex and stabilize a dynamic state of the spliceosome, respectively. Our findings thus show how multidimensional proteomic analysis of focused libraries of electrophilic compounds can expedite the discovery of chemical probes with site-specific functional effects on protein complexes in human cells.

Keywords
activity-based protein profiling; chemical probe; covalent; cysteine; proteasome; protein complexes; proteomics; size-exclusion chromatography; spliceosome.
Products