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Essential Role for Ubiquitin-Ubiquitin-Conjugating Enzyme Interaction in Ubiquitin Discharge from Cdc34 to Substrate

Saha, Anjanabha and Lewis, Steven and Kleiger, Gary and Kuhlman, Brian and Deshaies, Raymond J. (2011) Essential Role for Ubiquitin-Ubiquitin-Conjugating Enzyme Interaction in Ubiquitin Discharge from Cdc34 to Substrate. Molecular Cell, 42 (1). pp. 75-83. ISSN 1097-2765. PMCID PMC3091889. https://resolver.caltech.edu/CaltechAUTHORS:20110502-103732644

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Abstract

During ubiquitin conjugation, the thioester bond that links “donor” ubiquitin to ubiquitin-conjugating enzyme (E2) undergoes nucleophilic attack by the -amino group of an acceptor lysine, resulting in formation of an isopeptide bond. Models of ubiquitination have envisioned the donor ubiquitin to be a passive participant in this process. However, we show here that the I44A mutation in ubiquitin profoundly inhibits its ability to serve as a donor for ubiquitin chain initiation or elongation, but can be rescued by computationally predicted compensatory mutations in the E2 Cdc34. The donor defect of ubiquitin-I44A can be partially suppressed either by using a low pKa amine (hydroxylamine) as the acceptor or by performing reactions at higher pH, suggesting that the discharge defect arises in part due to inefficient deprotonation of the acceptor lysine. We propose that interaction between Cdc34 and the donor ubiquitin organizes the active site to promote efficient ubiquitination of substrate.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/j.molcel.2011.03.016DOIArticle
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3091889/PubMed CentralArticle
ORCID:
AuthorORCID
Deshaies, Raymond J.0000-0002-3671-9354
Additional Information:© 2011 Elsevier. Received: December 21, 2010; Revised: February 22, 2011; Accepted: March 24, 2011; Published: April 7, 2011. We thank P. Douglas Renfrew for assisting with generation of thioester torsion parameters. We also thank Michael Rape for generously sharing data prior to publication. A.S. was supported in part by a senior postdoctoral fellowship from the Leukemia and Lymphoma Society. R.J.D. is an HHMI Investigator, and this work was supported in part by HHMI and NIH grant GM065997.
Funders:
Funding AgencyGrant Number
Leukemia and Lymphoma Society UNSPECIFIED
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
NIHGM065997
Issue or Number:1
PubMed Central ID:PMC3091889
Record Number:CaltechAUTHORS:20110502-103732644
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20110502-103732644
Official Citation:Anjanabha Saha, Steven Lewis, Gary Kleiger, Brian Kuhlman, Raymond J. Deshaies, Essential Role for Ubiquitin-Ubiquitin-Conjugating Enzyme Interaction in Ubiquitin Discharge from Cdc34 to Substrate, Molecular Cell, Volume 42, Issue 1, 8 April 2011, Pages 75-83, ISSN 1097-2765, DOI: 10.1016/j.molcel.2011.03.016. (http://www.sciencedirect.com/science/article/B6WSR-52JV1GC-3/2/8b7599bee7ac53d2cbff25922c279a9f)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:23516
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:04 May 2011 18:38
Last Modified:17 Nov 2019 06:11

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