Systematic and Quantitative Assessment of the Ubiquitin-modified Proteome

Overview

Journal Mol Cell

Publisher Cell Press

Specialty Cell Biology

Date 2011 Sep 13

PMID 21906983

Citations 898

Authors

Woong Kim

Eric J Bennett

Edward L Huttlin

Ailan Guo

Jing Li

Anthony Possemato

Mathew E Sowa

Ramin Rad

John Rush

Michael J Comb

J Wade Harper

Steven P Gygi

Affiliations

Soon will be listed here.

Abstract

Despite the diverse biological pathways known to be regulated by ubiquitylation, global identification of substrates that are targeted for ubiquitylation has remained a challenge. To globally characterize the human ubiquitin-modified proteome (ubiquitinome), we utilized a monoclonal antibody that recognizes diglycine (diGly)-containing isopeptides following trypsin digestion. We identify ~19,000 diGly-modified lysine residues within ~5000 proteins. Using quantitative proteomics we monitored temporal changes in diGly site abundance in response to both proteasomal and translational inhibition, indicating both a dependence on ongoing translation to observe alterations in site abundance and distinct dynamics of individual modified lysines in response to proteasome inhibition. Further, we demonstrate that quantitative diGly proteomics can be utilized to identify substrates for cullin-RING ubiquitin ligases. Interrogation of the ubiquitinome allows for not only a quantitative assessment of alterations in protein homeostasis fidelity, but also identification of substrates for individual ubiquitin pathway enzymes.

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