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

Leiden Institute of Chemistry and Netherlands Proteomics Centre, Gorlaeus Laboratories, Einsteinweg 55, 2333 CC Leiden, The Netherlands. b.florea@chem.leidenuniv.nl

Chemistry & biology 2010 Aug 27

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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. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Citation

Bogdan I Florea, 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. Activity-based profiling reveals reactivity of the murine thymoproteasome-specific subunit beta5t. Chemistry & biology. 2010 Aug 27;17(8):795-801