2. Academic Validation
  3. Activity-based profiling reveals reactivity of the murine thymoproteasome-specific subunit beta5t

Activity-based profiling reveals reactivity of the murine thymoproteasome-specific subunit beta5t

  • Chem Biol. 2010 Aug 27;17(8):795-801. doi: 10.1016/j.chembiol.2010.05.027.
Bogdan I Florea 1,  Martijn Verdoes,  Nan Li,  Wouter A van der Linden,  Paul P Geurink,  Hans van den Elst,  Tanja Hofmann,  Arnoud de Ru,  Peter A van Veelen,  Keiji Tanaka,  Katsuhiro Sasaki,  Shigeo Murata,  Hans den Dulk,  Jaap Brouwer,  Ferry A Ossendorp,  Alexei F Kisselev,  Herman S Overkleeft
Affiliations

Affiliation

  • 1 Leiden Institute of Chemistry and Netherlands Proteomics Centre, Gorlaeus Laboratories, Einsteinweg 55, 2333 CC Leiden, The Netherlands. [email protected]
PMID: 20797608 DOI: 10.1016/j.chembiol.2010.05.027
Abstract

Epithelial cells of the thymus cortex express a unique Proteasome particle involved in positive T cell selection. This thymoproteasome contains the recently discovered beta5t subunit that has an uncharted activity, if any. We synthesized fluorescent epoxomicin probes that were used in a chemical proteomics approach, entailing activity-based profiling, affinity purification, and LC-MS identification, to demonstrate that the beta5t subunit is catalytically active in the murine thymus. A panel of established Proteasome inhibitors showed that the broad-spectrum inhibitor epoxomicin blocks the beta5t activity and that the subunit-specific antagonists bortezomib and NC005 do not inhibit beta5t. We show that beta5t has a substrate preference distinct from beta5/beta5i that might explain how the thymoproteasome generates the MHC class I peptide repertoire needed for positive T cell selection.

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