Deconvolution of substrate processing by the 26S proteasome reveals a selective kinetic gateway to degradation
The 26S proteasome is the principle macromolecular machine responsible for protein degradation in eukaryotes. However, little is known about the detailed kinetics and coordination of the underlying substrate-processing steps of the proteasome, and their correlation with observed conformational states. Here, we used reconstituted 26S proteasomes with unnatural amino acid-attached fluorophores in a series of FRET and anisotropy-based assays to probe substrate-proteasome interactions, the individual steps of the processing pathway, and the conformational state of the proteasome itself. We develop a complete kinetic picture of proteasomal degradation, which reveals that the engagement steps prior to substrate commitment are fast relative to subsequent deubiquitination, translocation and unfolding. Furthermore, we find that non-ideal substrates are rapidly rejected by the proteasome, which thus employs a kinetic proofreading mechanism to ensure degradation fidelity and substrate prioritization.
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Jared AM Bard (add twitter)
Charlene Bashore (add twitter)
Ken C Dong (add twitter)
Andreas Martin (add twitter)
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Biochemistry

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06/30/18 07:47AM
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WordenEvan: long time in the making. Complete deconvolution of substrate processing by the 26S proteasome. All kinetic steps revealed! Great work by my old lab @proteazomnomnom https://t.co/eMIGXfKdjK
AlanLHutchison: Excited to read @jabard89 new @biorxivpreprint "Deconvolution of substrate processing by the 26S proteasome reveals a selective kinetic gateway to degradation". Up and coming scientist soon joining the lab of @dallandrummond as a @bsdpostdoc! https://t.co/WTZG6y7hBu
proteazomnomnom: The preprint of our study on the kinetics of substrate processing by the 26S #proteasome is out: https://t.co/1wyK4wh5ym https://t.co/YJgs19b4xZ
biorxivpreprint: Deconvolution of substrate processing by the 26S proteasome reveals a selective kinetic gateway to degradation https://t.co/1JCZWyxImZ #bioRxiv
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