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Quantitative Determination of MAR Hydrolase Residue Specificity In Vitro by Tandem Mass Spectrometry

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ADP-ribosylation and NAD+ Utilizing Enzymes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1813))

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Abstract

ADP-ribosylation is a posttranslational modification that involves the conjugation of monomers and polymers of the small molecule ADP-ribose onto amino acid side chains. A family of ADP-ribosyltransferases catalyzes the transfer of the ADP-ribose moiety of nicotinamide adenine dinucleotide (NAD+) onto a variety of amino acid side chains including aspartate, glutamate, lysine, arginine, cysteine, and serine. The monomeric form of the modification mono(ADP-ribosyl)ation (MARylation) is reversed by a number of enzymes including a family of MacroD-type macrodomain-containing mono(ADP-ribose) (MAR) hydrolases. Though it has been inferred from various chemical tests that these enzymes have specificity for MARylated aspartate and glutamate residues in vitro, the amino acid and site specificity of different family members are often not unambiguously defined. Here we describe a mass spectrometry-based assay to determine the site specificity of MAR hydrolases in vitro.

The original version of this chapter was revised. A correction to this chapter can be found at https://doi.org/10.1007/978-1-4939-8588-3_27

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Change history

  • 16 November 2018

    The funding information was omitted from the original Chapters 19 and 21. The below text has been added to these chapters respectively.

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Acknowledgments

This work was supported by a Johns Hopkins Catalyst Award (AKLL) and research grants from the Johns Hopkins University School of Medicine Sherrilyn and Ken Fisher Center for Environmental Infectious Disease (AKLL). The proteomics work was also in part supported by R01GM104135S1 (AKLL and RLM), S10OD021502 (SEO), and T32CA009110 (RLM) from the U.S. National Institutes of Health, and the American Cancer Society Research Scholar Award 129539-RSG-16-062-01-RMC (AKLL).

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Correspondence to Anthony K. L. Leung .

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McPherson, R.L., Ong, SE., Leung, A.K.L. (2018). Quantitative Determination of MAR Hydrolase Residue Specificity In Vitro by Tandem Mass Spectrometry. In: Chang, P. (eds) ADP-ribosylation and NAD+ Utilizing Enzymes. Methods in Molecular Biology, vol 1813. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-8588-3_19

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  • DOI: https://doi.org/10.1007/978-1-4939-8588-3_19

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-8587-6

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