Labeling and quantifying sites of protein palmitoylation

Biotechniques. 2004 Feb;36(2):276-85. doi: 10.2144/04362RR02.


As a reversible posttranslational modification, protein palmitoylation has the potential to regulate the trafficking and function of a variety of proteins. However, the extent, function, and dynamic nature of palmitoylation are poorly resolved because of limitations in assay methods. Here, we introduce methods where hydroxylamine-mediated cleavage of the palmitoyl-thioester bond generates a free sulfhydryl, which can then be specifically labeled with sulfhydryl-reactive reagents. This methodology is more sensitive and allows for quantitative estimates of palmitoylation. Unlike other techniques used to assay posttranslational modifications, the techniques we have developed can label all sites of modification with a variety of probes, radiolabeled or nonradioactive, and can be used to assay the palmitoylation of proteins expressed in vivo in brain or other tissues.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Binding Sites
  • Biotinylation
  • Blotting, Western / methods
  • Cell Line
  • Chickens
  • Ethylmaleimide / metabolism
  • Genetic Vectors
  • Humans
  • Membrane Proteins / metabolism
  • Mice
  • Nerve Tissue Proteins / metabolism
  • Palmitates / analysis
  • Palmitates / metabolism*
  • Protein Binding
  • Proteins / analysis
  • Proteins / metabolism*
  • Staining and Labeling / methods*
  • Synaptosomal-Associated Protein 25
  • Transfection
  • Tritium / metabolism


  • Membrane Proteins
  • Nerve Tissue Proteins
  • Palmitates
  • Proteins
  • SNAP25 protein, human
  • Snap25 protein, mouse
  • Synaptosomal-Associated Protein 25
  • Tritium
  • Ethylmaleimide