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Proteome-wide identification of in vivo ADP-ribose acceptor sites by liquid chromatography–tandem mass spectrometry


Larsen, Sara C; Leutert, Mario; Bilan, Vera; Martello, Rita; Jungmichel, Stephanie; Young, Clifford; Hottiger, Michael O; Nielsen, Michael L (2017). Proteome-wide identification of in vivo ADP-ribose acceptor sites by liquid chromatography–tandem mass spectrometry. In: Tulin, Alexei V. Poly(ADP-Ribose) Polymerase. Switzerland: Springer, 149-162.

Abstract

ADP-ribosylation is a posttranslational modification (PTM) that affects a variety of cellular processes. In recent years, mass spectrometry (MS)-based proteomics has become a valuable tool for studying ADP-ribosylation. However, studying this PTM in vivo in an unbiased and sensitive manner has remained a difficult challenge. Here, we describe a detailed protocol for unbiased analysis of ADP-ribosylated proteins and their ADP-ribose acceptor sites under physiological conditions. The method relies on the enrichment of mono-ADP-ribosylated peptides using the macrodomain Af1521 in combination with liquid chromatography-high-resolution tandem MS (LC-MS/MS). The 5-day protocol explains the step-by-step enrichment and identification of ADP-ribosylated peptides from cell culture stage all the way through to data processing using the MaxQuant software suite.

Abstract

ADP-ribosylation is a posttranslational modification (PTM) that affects a variety of cellular processes. In recent years, mass spectrometry (MS)-based proteomics has become a valuable tool for studying ADP-ribosylation. However, studying this PTM in vivo in an unbiased and sensitive manner has remained a difficult challenge. Here, we describe a detailed protocol for unbiased analysis of ADP-ribosylated proteins and their ADP-ribose acceptor sites under physiological conditions. The method relies on the enrichment of mono-ADP-ribosylated peptides using the macrodomain Af1521 in combination with liquid chromatography-high-resolution tandem MS (LC-MS/MS). The 5-day protocol explains the step-by-step enrichment and identification of ADP-ribosylated peptides from cell culture stage all the way through to data processing using the MaxQuant software suite.

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Additional indexing

Item Type:Book Section, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Department of Molecular Mechanisms of Disease
07 Faculty of Science > Department of Molecular Mechanisms of Disease
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:July 2017
Deposited On:09 Aug 2017 10:18
Last Modified:21 Sep 2017 10:19
Publisher:Springer
Series Name:Methods in Molecular Biology
Number:1608
ISSN:1064-3745
ISBN:978-1-4939-6992-0
Publisher DOI:https://doi.org/10.1007/978-1-4939-6993-7_11
Related URLs:http://www.recherche-portal.ch/ZAD:default_scope:ebi01_prod000916990 (Library Catalogue)
PubMed ID:28695509

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