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Detection of Protein S-Acylation using Acyl-Resin Assisted Capture
- Source :
- J Vis Exp
- Publication Year :
- 2020
- Publisher :
- MyJove Corporation, 2020.
-
Abstract
- Protein S-acylation, also referred to as S-palmitoylation, is a reversible post-translational modification of cysteine residues with long-chain fatty acids via a labile thioester bond. S-acylation, which is emerging as a widespread regulatory mechanism, can modulate almost all aspects of the biological activity of proteins, from complex formation to protein trafficking and protein stability. The recent progress in understanding of the biological function of protein S-acylation was achieved largely due to the development of novel biochemical tools allowing robust and sensitive detection of protein S-acylation in a variety of biological samples. Here, we describe acyl resin-assisted capture (Acyl-RAC), a recently developed method based on selective capture of endogenously S-acylated proteins by thiol-reactive Sepharose beads. Compared to existing approaches, Acyl-RAC requires fewer steps and can yield more reliable results when coupled with mass spectrometry for identification of novel S-acylation targets. A major limitation in this technique is the lack of ability to discriminate between fatty acid species attached to cysteines via the same thioester bond.
- Subjects :
- 0301 basic medicine
chemistry.chemical_classification
General Immunology and Microbiology
Acylation
General Chemical Engineering
General Neuroscience
Fatty acid
S-acylation
Biological activity
Thioester
Article
General Biochemistry, Genetics and Molecular Biology
Protein S
Sepharose
03 medical and health sciences
030104 developmental biology
0302 clinical medicine
Palmitoylation
chemistry
Biochemistry
lipids (amino acids, peptides, and proteins)
030217 neurology & neurosurgery
Cysteine
Subjects
Details
- ISSN :
- 1940087X
- Database :
- OpenAIRE
- Journal :
- Journal of Visualized Experiments
- Accession number :
- edsair.doi.dedup.....a4874e899f76750b753a2f39ba547121