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Mechanistic Dissection of the Cop9 Signalosome’s Deneddylation Activity on Cullin-RING Ligases


Mosadeghi, Ruzbeh (2015) Mechanistic Dissection of the Cop9 Signalosome’s Deneddylation Activity on Cullin-RING Ligases. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z9P26W3G.


We set out to understand the precise mechanisms that regulate the activation and deactivation of Cullin-RING Ligases (CRLs). While a great deal of work has already gone into identifying the players involved in these pathways and the cellular consequences associated with the loss of each, the biochemical mechanisms regulating these steps have remained elusive. In this work we sought to gain a better understanding of the mechanisms behind these steps by teasing apart specific their biochemical reactions. By measuring the individual microscopic rate constants of the reactions we have shed light on both the proper sequence of events in the regulation of CRLs as well as how they are in fact controlled.

Prior to this work, it was believed that CSN deactivated CRLs by binding them and enzymatically removing the activating post-translation modification Nedd8. It was believed that CSN could not bind to CRLs while they were active due to the steric hindrance by the CRL substrates, and that they would remain bound to deneddylated CRLs as a sequestering agent until a new substrate could displace it. We now have some insight that substrates themselves cannot inhibit CSN very well, but that the active ubiquitination by an E2 enzyme precludes CSN binding and activity. When the substrate for a CRL becomes depleted, CSN then binds to the CRL in a low affinity, low activity conformation. This triggers a conformational change that pulls the autoinhibitory Ins-1 loop away from the active site in the catalytic subunit Csn5, resulting in a large increase in affinity and cleavage of the isopeptide bond between CRLs and Nedd8. Upon dissociation of Nedd8, CSN rapidly returns to the low affinity state and dissociates from the CRL, allowing it reenter its activation cycle.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:Cullin-RING Ligase, CRL, Cop9 Signalosome, CSN, Nedd8
Degree Grantor:California Institute of Technology
Division:Biology and Biological Engineering
Major Option:Biology
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Deshaies, Raymond Joseph
Thesis Committee:
  • Chan, David C. (chair)
  • Mazmanian, Sarkis K.
  • Shan, Shu-ou
  • Deshaies, Raymond Joseph
Defense Date:3 April 2015
Record Number:CaltechTHESIS:05312015-231120184
Persistent URL:
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:8951
Deposited By: Ruzbeh Mosadeghi
Deposited On:06 Oct 2016 23:14
Last Modified:04 Oct 2019 00:08

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