| Autor: |
Buffa V; Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, Italy.; Integrare UMR_S951 Genethon, Inserm, University of Evry, Université Paris Saclay Genethon, 91000 Evry, France., Adamo G; Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, Italy., Picciotto S; Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, Italy., Bongiovanni A; Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, Italy., Romancino DP; Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, Italy. |
| Abstrakt: |
Protein S-palmitoylation is a reversible post-translational lipidation in which palmitic acid (16:0) is added to protein cysteine residue by a covalent thioester bond. This modification plays an active role in membrane targeting of soluble proteins, protein-protein interaction, protein trafficking, and subcellular localization. Moreover, palmitoylation is related to different diseases, such as neurodegenerative pathologies, cancer, and developmental defects. The aim of this research is to provide a straightforward and sensitive procedure to detect protein palmitoylation based on Acyl Biotin Exchange (ABE) chemistry. Our protocol setup consists of co-immunoprecipitation of native proteins (i.e., CD63), followed by the direct detection of palmitoylation on proteins immobilized on polyvinylidene difluoride (PVDF) membranes. With respect to the conventional ABE-based protocol, we optimized and validated a rapid semi-quantitative assay that is shown to be significantly more sensitive and highly reproducible. |
