Quantification of mitochondrial S-Nitrosylation by CysTMT6 switch assay

Christopher I. Murray, Hea Seung Chung, Helge Uhrigshardt, Jennifer E. Van Eyk

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

S-nitrosylation (SNO) is an important oxidative posttranslational modification in the regulation of cardiac mitochondria. SNO modification of several mitochondrial proteins has been associated with cardiac preconditioning and improved cell survival following ischemia/reperfusion injury. Due to their labile nature, SNO modifications are challenging to study using traditional biochemical techniques; particularly, the identification of individual modified cysteine residues. Here, we describe the details of the cysTMT 6 switch assay, a variation of the classic biotin switch protocol. The cysTMT 6 reagent provides a simplified and powerful approach to SNO detection by combining unambiguous identification of the modified cysteine residue and relative quantification of up to six samples by mass spectrometry analysis.

Original languageEnglish (US)
Title of host publicationHeart Proteomics
Subtitle of host publicationMethods and Protocols
PublisherHumana Press Inc.
Pages169-179
Number of pages11
ISBN (Print)9781627033855
DOIs
StatePublished - 2013

Publication series

NameMethods in Molecular Biology
Volume1005
ISSN (Print)1064-3745

Keywords

  • Biotin switch assay
  • Cysteine-reactive tandem mass tags
  • Mitochondria
  • Oxidative modification
  • Quantitation
  • S-nitrosylation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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