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Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells
Identification of protein interactors is fundamental to understanding their functions. Here, we describe a modified protocol for tandem affinity purification coupled with mass spectrometry (TAP/MS), which includes two-step purification. We detail the S-, 2×FLAG-, and Streptavidin-Binding Peptide (SB...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9304681/ https://www.ncbi.nlm.nih.gov/pubmed/35874475 http://dx.doi.org/10.1016/j.xpro.2022.101569 |
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author | Bian, Weixiang Jiang, Hua Feng, Shan Chen, Junjie Wang, Wenqi Li, Xu |
author_facet | Bian, Weixiang Jiang, Hua Feng, Shan Chen, Junjie Wang, Wenqi Li, Xu |
author_sort | Bian, Weixiang |
collection | PubMed |
description | Identification of protein interactors is fundamental to understanding their functions. Here, we describe a modified protocol for tandem affinity purification coupled with mass spectrometry (TAP/MS), which includes two-step purification. We detail the S-, 2×FLAG-, and Streptavidin-Binding Peptide (SBP)- tandem tags (SFB-tag) system for protein purification. This protocol can be used to identify protein interactors and establish a high-confidence protein–protein interaction network based on computational models. This is particularly useful for identifying bona fide interacting proteins for subsequent functional studies. For complete details on the use and execution of this protocol, please refer to Bian et al. (2021). |
format | Online Article Text |
id | pubmed-9304681 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-93046812022-07-23 Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells Bian, Weixiang Jiang, Hua Feng, Shan Chen, Junjie Wang, Wenqi Li, Xu STAR Protoc Protocol Identification of protein interactors is fundamental to understanding their functions. Here, we describe a modified protocol for tandem affinity purification coupled with mass spectrometry (TAP/MS), which includes two-step purification. We detail the S-, 2×FLAG-, and Streptavidin-Binding Peptide (SBP)- tandem tags (SFB-tag) system for protein purification. This protocol can be used to identify protein interactors and establish a high-confidence protein–protein interaction network based on computational models. This is particularly useful for identifying bona fide interacting proteins for subsequent functional studies. For complete details on the use and execution of this protocol, please refer to Bian et al. (2021). Elsevier 2022-07-19 /pmc/articles/PMC9304681/ /pubmed/35874475 http://dx.doi.org/10.1016/j.xpro.2022.101569 Text en © 2022 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). |
spellingShingle | Protocol Bian, Weixiang Jiang, Hua Feng, Shan Chen, Junjie Wang, Wenqi Li, Xu Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title | Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title_full | Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title_fullStr | Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title_full_unstemmed | Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title_short | Protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
title_sort | protocol for establishing a protein-protein interaction network using tandem affinity purification followed by mass spectrometry in mammalian cells |
topic | Protocol |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9304681/ https://www.ncbi.nlm.nih.gov/pubmed/35874475 http://dx.doi.org/10.1016/j.xpro.2022.101569 |
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