Refined Preparation and Use of Anti-diglycine Remnant (K-ε-GG) Antibody Enables Routine Quantification of 10,000s of Ubiquitination Sites in Single Proteomics Experiments

Overview

Journal Mol Cell Proteomics

Specialties Biochemistry
Cell Biology
Molecular Biology

Date 2012 Dec 26

PMID 23266961

Citations 174

Authors

Namrata D Udeshi

Tanya Svinkina

Philipp Mertins

Eric Kuhn

D R Mani

Jana W Qiao

Steven A Carr

Affiliations

Soon will be listed here.

Abstract

Detection of endogenous ubiquitination sites by mass spectrometry has dramatically improved with the commercialization of anti-di-glycine remnant (K-ε-GG) antibodies. Here, we describe a number of improvements to the K-ε-GG enrichment workflow, including optimized antibody and peptide input requirements, antibody cross-linking, and improved off-line fractionation prior to enrichment. This refined and practical workflow enables routine identification and quantification of ∼20,000 distinct endogenous ubiquitination sites in a single SILAC experiment using moderate amounts of protein input.

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