Identification of S-Nitrosothiols by the Sequential Cysteine Blocking Technique.
Here, we describe a procedure for the identification of S-nitrosothiols that has been used in our laboratory to study the roles of protein S-nitrosylation in the immune responses of Arabidopsis thaliana and other organisms. It employs a modified version of the biotin-switch technique, which we termed the sequential cysteine blocking technique, encompassing the sequential redox-blocking of recombinant proteins followed by LC-MS/MS analysis.
Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, King's Buildings, Edinburgh, EH9 3JH, UK.
Rafael A
RA
Homem
Synthetic and Systems Biology, School of Biological Sciences, University of Edinburgh, King's Buildings, Edinburgh, EH9 3JH, UK.
Thierry
T
Le Bihan
Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, King's Buildings, Edinburgh, EH9 3JH, UK.
Manda
M
Yu
Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, King's Buildings, Edinburgh, EH9 3JH, UK. gloake@ed.ac.uk.|Synthetic and Systems Biology, School of Biological Sciences, University of Edinburgh, King's Buildings, Edinburgh, EH9 3JH, UK. gloake@ed.ac.uk.
Gary J
GJ
Loake
D017360:Arabidopsis / Q000378:metabolism; D002853:Chromatography, Liquid; D003545:Cysteine / Q000378:metabolism*; D026403:S-Nitrosothiols / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
D026403:S-Nitrosothiols; D003545:Cysteine
Iodoacetamide; LC–MS/MS; Label-free quantitation; Redox-based posttranslational modification; S-nitrosothiols; S-nitrosylation; Sequential cysteine blocking