Chapter title |
Proteome-Wide Identification of In Vivo ADP-Ribose Acceptor Sites by Liquid Chromatography–Tandem Mass Spectrometry
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Chapter number | 11 |
Book title |
Poly(ADP-Ribose) Polymerase
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Published in |
Methods in molecular biology, July 2017
|
DOI | 10.1007/978-1-4939-6993-7_11 |
Pubmed ID | |
Book ISBNs |
978-1-4939-6992-0, 978-1-4939-6993-7
|
Authors |
Sara C. Larsen, Mario Leutert, Vera Bilan, Rita Martello, Stephanie Jungmichel, Clifford Young, Michael O. Hottiger, Michael L. Nielsen, Larsen, Sara C., Leutert, Mario, Bilan, Vera, Martello, Rita, Jungmichel, Stephanie, Young, Clifford, Hottiger, Michael O., Nielsen, Michael L. |
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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