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Identification of ADP-Ribose acceptor sites on in vitro modified proteins by liquid chromatography–tandem mass spectrometry


Leutert, Mario; Bilan, Vera; Gehrig, Peter; Hottiger, Michael O (2017). Identification of ADP-Ribose acceptor sites on in vitro modified proteins by liquid chromatography–tandem mass spectrometry. Methods in Molecular Biology, 1608:137-148.

Abstract

Protein ADP-ribosylation is a covalent, reversible posttranslational modification (PTM) catalyzed by ADP-ribosyltransferases (ARTs). Proteins can be either mono- or poly-ADP-ribosylated under a variety of physiological and pathological conditions. To understand the functional contribution of protein ADP-ribosylation to normal and disease/stress states, modified protein and corresponding ADP-ribose acceptor site identification is crucial. Since ADP-ribosylation is a transient and relatively low abundant PTM, systematic and accurate identification of ADP-ribose acceptor sites has only recently become feasible. This is due to the development of specific ADP-ribosylated protein/peptide enrichment methodologies, as well as technical advances in high-accuracy liquid chromatography-tandem mass spectrometry (LC-MS/MS). The standardized protocol described here allows the identification of ADP-ribose acceptor sites in in vitro ADP-ribosylated proteins and will, thus, contribute to the functional characterization of this important PTM.

Abstract

Protein ADP-ribosylation is a covalent, reversible posttranslational modification (PTM) catalyzed by ADP-ribosyltransferases (ARTs). Proteins can be either mono- or poly-ADP-ribosylated under a variety of physiological and pathological conditions. To understand the functional contribution of protein ADP-ribosylation to normal and disease/stress states, modified protein and corresponding ADP-ribose acceptor site identification is crucial. Since ADP-ribosylation is a transient and relatively low abundant PTM, systematic and accurate identification of ADP-ribose acceptor sites has only recently become feasible. This is due to the development of specific ADP-ribosylated protein/peptide enrichment methodologies, as well as technical advances in high-accuracy liquid chromatography-tandem mass spectrometry (LC-MS/MS). The standardized protocol described here allows the identification of ADP-ribose acceptor sites in in vitro ADP-ribosylated proteins and will, thus, contribute to the functional characterization of this important PTM.

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Additional indexing

Item Type:Journal Article, 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
Uncontrolled Keywords:ADP-ribosylation; ADP-ribosylome; ARTD; Mass spectrometry; PARG; PARP; Phosphoenrichment; Ti4+-IMAC enrichment
Language:English
Date:2017
Deposited On:08 Aug 2017 16:23
Last Modified:09 Dec 2017 01:42
Publisher:Springer
ISSN:1064-3745
ISBN:978-1-4939-6992-0
Publisher DOI:https://doi.org/10.1007/978-1-4939-6993-7_10
PubMed ID:28695508

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