A Quantitative Chemical Proteomics Approach for Site-specific Stoichiometry Analysis of Ubiquitination

Yunan Li, Jonathan Evers, Ang Luo, Luke Erber, Zachary Postler, Yue Chen

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

Stoichiometric analysis of post-translational modifications is an emerging strategy for absolute quantification of the fractional abundance of the modification. Herein, a quantitative chemical proteomic workflow for stoichiometric analysis of ubiquitination is reported, named isotopically balanced quantification of ubiquitination (IBAQ-Ub). The strategy utilizes a new amine-reactive chemical tag (AcGG-NHS) that is structurally homologous to the GG remnant of ubiquitin on modified lysine after trypsin cleavage and therefore enables the generation of structurally identical peptides from ubiquitinated and unmodified lysine residues following trypsin digestion and secondary stable isotopic labeling. The strategy is highly robust, sensitive, and accurate with a wide dynamic range using either protein standards or complex cell lysates. Thus, this work provides an efficient chemical proteomics tool for quantitative stoichiometric analysis of ubiquitination signaling pathways.

Original languageEnglish (US)
Pages (from-to)537-541
Number of pages5
JournalAngewandte Chemie - International Edition
Volume58
Issue number2
DOIs
StatePublished - Jan 8 2019

Bibliographical note

Funding Information:
We greatly appreciate the Center for Mass Spectrometry and Proteomics and the Masonic Cancer Center at the University of Minnesota for LC-MS access and WuXi Apptec. Co. for the technical support on chemical synthesis. This work was supported by the research start-up fund to Y.C. from the University of Minnesota and National Science Foundation (CHE-1753154 to Y.C.).

Keywords

  • IBAQ-Ub
  • proteasome inhibition
  • quantitative proteomics
  • stoichiometry
  • ubiquitination

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