In vivo measurement of the acetylation state of sirtuin substrates as a proxy for sirtuin activity

John Dominy, Pere Puigserver, Carles Cantó

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Evaluating the precise catalytic activity of sirtuin proteins in vivo is a challenging endeavor. Enzymological methods, including those employed in commercially available kits, require the isolation of immunopurified protein from cells or tissues, which can perturb regulatory protein-protein interactions as well as remove the enzyme from the reaction-altering effects of intracellular NAD+, nicotinamide, and O-acetyl-ADP ribose concentrations. As such, the measurement of the steady state acetylation status of select sirtuin substrates in vivo remains an important tool for evaluating changes in sirtuin activity. Here, we describe how to perform the analysis of the acetylation status of key SIRT1 and SIRT3 targets in rodent tissues and cultured cells.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
Pages217-237
Number of pages21
Volume1077
DOIs
StatePublished - 2013
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume1077
ISSN (Print)10643745

Keywords

  • Acetylation
  • Homogenization
  • Immunoprecipitation
  • Ndufa9
  • PGC-1α
  • Sirtuins
  • Transfection
  • Western blot

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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