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Assays for NAD+-Dependent Reactions and NAD+ Metabolites

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ADP-ribosylation and NAD+ Utilizing Enzymes

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

Abstract

Nicotinamide adenine dinucleotide (NAD+) is an essential redox cofactor and signaling molecule that controls the activity of enzymes involved in metabolism, DNA repair, and cellular survival, such as the PARPs, CD38, and the sirtuins. Here, we describe three methods for measuring the activity of these enzymes: the etheno-NAD+ assay measures NAD+ hydrolase activity using an NAD+ analog to produce a fluorescent product that is measured in real time; the PNC1 assay converts a native product of NAD+ hydrolysis, nicotinamide, into a quantitative fluorescent readout; and liquid chromatography tandem mass spectrometry (LC-MS/MS) is used to characterize the entire NAD+ metabolome in a sample. These methods will enable new insights into the roles that NAD+ and the enzymes that utilize it play in health and disease.

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Acknowledgments

N.B. is the recipient of the Australian Research Council Discovery Early Career Research Award at the University of New South Wales, Sydney, Australia.

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Authors and Affiliations

  1. Department of Genetics, Paul F. Glenn Center for the Biology of Aging, Harvard Medical School, Boston, MA, USA

    Michael B. Schultz, Yuancheng Lu & David A. Sinclair

  2. Centre for Healthy Brain Ageing, School of Psychiatry, The University of New South Wales, Sydney, NSW, Australia

    Nady Braidy

  3. Department of Pharmacology, School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia

    David A. Sinclair

Authors
  1. Michael B. Schultz
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  2. Yuancheng Lu
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  3. Nady Braidy
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  4. David A. Sinclair
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Corresponding author

Correspondence to David A. Sinclair .

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Editors and Affiliations

  1. Department of Biology, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA

    Paul Chang

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Schultz, M.B., Lu, Y., Braidy, N., Sinclair, D.A. (2018). Assays for NAD+-Dependent Reactions and NAD+ Metabolites. 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_6

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  • DOI: https://doi.org/10.1007/978-1-4939-8588-3_6

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-8587-6

  • Online ISBN: 978-1-4939-8588-3

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