A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels
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,...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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MDPI AG
2023-03-01
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| Series: | Membranes |
| Subjects: | |
| Online Access: | https://www.mdpi.com/2077-0375/13/3/361 |
| _version_ | 1827748778864541696 |
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| author | Valentina Buffa Giorgia Adamo Sabrina Picciotto Antonella Bongiovanni Daniele P. Romancino |
| author_facet | Valentina Buffa Giorgia Adamo Sabrina Picciotto Antonella Bongiovanni Daniele P. Romancino |
| author_sort | Valentina Buffa |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-03-11T06:11:37Z |
| format | Article |
| id | doaj.art-c24370bb2e8c4b508aeed30441849a49 |
| institution | Directory Open Access Journal |
| issn | 2077-0375 |
| language | English |
| last_indexed | 2024-03-11T06:11:37Z |
| publishDate | 2023-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Membranes |
| spelling | doaj.art-c24370bb2e8c4b508aeed30441849a492023-11-17T12:35:29ZengMDPI AGMembranes2077-03752023-03-0113336110.3390/membranes13030361A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation LevelsValentina Buffa0Giorgia Adamo1Sabrina Picciotto2Antonella Bongiovanni3Daniele P. Romancino4Institute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, ItalyInstitute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, ItalyInstitute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, ItalyInstitute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, ItalyInstitute for Research and Biomedical Innovation (IRIB), National Research Council (CNR), Via Ugo La Malfa, 153-90146 Palermo, ItalyProtein 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.https://www.mdpi.com/2077-0375/13/3/361membrane proteinpost-translational lipidationS-palmitoylationacyl biotin exchange (ABE) |
| spellingShingle | Valentina Buffa Giorgia Adamo Sabrina Picciotto Antonella Bongiovanni Daniele P. Romancino A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels Membranes membrane protein post-translational lipidation S-palmitoylation acyl biotin exchange (ABE) |
| title | A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels |
| title_full | A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels |
| title_fullStr | A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels |
| title_full_unstemmed | A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels |
| title_short | A Simple, Semi-Quantitative Acyl Biotin Exchange-Based Method to Detect Protein S-Palmitoylation Levels |
| title_sort | simple semi quantitative acyl biotin exchange based method to detect protein s palmitoylation levels |
| topic | membrane protein post-translational lipidation S-palmitoylation acyl biotin exchange (ABE) |
| url | https://www.mdpi.com/2077-0375/13/3/361 |
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