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bioRxiv · 10.1101/2024.11.07.622408

Salt-dependent non-catalytic allostery of human USP14-regulated 26S proteasome

Zou, S.·Zhang, S.·Zhao, L.·Mao, Y.
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Abstract

USP14, a deubiquitylating enzyme, regulates 26S proteasome function both catalytically and non-catalytically through multiple checkpoints. How USP14 non-catalytically regulates the proteasome activity remains elusive. Here, we combined genetic engineering and cryo-EM to disentangle how USP14 non-catalytically regulates proteasome activity in a salt-dependent manner. By solving 16 high-resolution cryo-EM structures of substrate-engaged human 26S proteasome complexed with a catalytically deficient mutant USP14, we demonstrate that USP14s non-catalytic activity alone can induce parallel pathways of proteasome state transitions, leading to opposing substrate fates. The USP14 mutant allosterically reprograms the AAA-ATPase motor, inducing novel substrate-engaged conformations and filling major gaps in understanding asymmetric ATP-hydrolysis cycling around the ATPase ring. Time-resolved cryo-EM and functional analysis reveal that sodium or potassium promotes substrate-engaged pathways and suppresses USP14 activity for enhanced degradation, uncovering another layer of complexity in proteasome regulation.

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Zou, S., Zhang, S., Zhao, L., Mao, Y.. 2024-11-07. Salt-dependent non-catalytic allostery of human USP14-regulated 26S proteasome. https://doi.org/10.1101/2024.11.07.622408

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