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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...

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Detalles Bibliográficos
Autores principales: Bian, Weixiang, Jiang, Hua, Feng, Shan, Chen, Junjie, Wang, Wenqi, Li, Xu
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2022
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).
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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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