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Assays for NAD+-Dependent Reactions and NAD+ Metabolites

  • Michael B. Schultz
  • Yuancheng Lu
  • Nady Braidy
  • David A. Sinclair
Protocol
Part of the Methods in Molecular Biology book series (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.

Key words

Aging NAD+ nicotinamide ADP-ribose PARP ARTD CD38 BST1 Sirtuin Metabolomics Mass spectrometry Metabolism Epigenetics HDAC 

Notes

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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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Michael B. Schultz
    • 1
  • Yuancheng Lu
    • 1
  • Nady Braidy
    • 2
  • David A. Sinclair
    • 1
    • 3
  1. 1.Department of Genetics, Paul F. Glenn Center for the Biology of AgingHarvard Medical SchoolBostonUSA
  2. 2.Centre for Healthy Brain Ageing, School of PsychiatryThe University of New South WalesSydneyAustralia
  3. 3.Department of Pharmacology, School of Medical SciencesThe University of New South WalesSydneyAustralia

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