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Methods Mol Biol. 2018;1813:205-213. doi: 10.1007/978-1-4939-8588-3_13.

Mono-ADP-Ribosylhydrolase Assays.

Author information

1
Department of Molecular Mechanisms of Disease, University of Zurich, Zurich, Switzerland.
2
Molecular Life Science PhD Program of the Life Science Zurich Graduate School, Zurich, Switzerland.
3
Department of Molecular Mechanisms of Disease, University of Zurich, Zurich, Switzerland. michael.hottiger@dmmd.uzh.ch.

Abstract

Despite substantial progress in ADP-ribosylation research in recent years, the identification of ADP-ribosylated proteins, their ADP-ribose acceptors sites, and the respective writers and erasers remains challenging. The use of recently developed mass spectrometric methods helps to further characterize the ADP-ribosylome and its regulatory enzymes under different conditions and in different cell types. Validation of these findings may be achieved by in vitro assays for the respective enzymes. In the below method, we describe how recombinant ADP-ribosylated proteins are demodified in vitro with mono-ADP-ribosylhydrolases of choice to elucidate substrate and potentially also site specificity of these enzymes.

KEYWORDS:

ARH1; ARH3; C6ORF130; De-ADP-ribosylation assay; MACROD1; MACROD2; Macrodomain; Mono-ADP-ribosylhydrolases; OARD1; PARG; TARG

PMID:
30097869
DOI:
10.1007/978-1-4939-8588-3_13
[Indexed for MEDLINE]

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