A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells

Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteomics to c...

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Detalles Bibliográficos
Autores principales: Zhu, Junji, Xiao, Wuhan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2021
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8599494/
https://www.ncbi.nlm.nih.gov/pubmed/34820639
http://dx.doi.org/10.1016/j.xpro.2021.100962
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author Zhu, Junji
Xiao, Wuhan
author_facet Zhu, Junji
Xiao, Wuhan
author_sort Zhu, Junji
collection PubMed
description Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteomics to compare stimulation-induced interactomes between wild-type and CRISPR-Cas9-mediated knockout cells. The candidates of interest are identified using bioinformatic analyses and verified by biochemical approaches. This protocol is highly versatile and applies to a variety of cells and different types of stimulation. For complete details on the use and execution of this protocol, please refer to (Zhu et al., 2021).
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spelling pubmed-85994942021-11-23 A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells Zhu, Junji Xiao, Wuhan STAR Protoc Protocol Some protein-protein interactions are induced by different kinds of stimulation and are dependent on specific genes. To identify these interaction partners, we present a protocol which utilizes affinity purification of Flag-tagged protein complexes followed by mass-spectrometry-based proteomics to compare stimulation-induced interactomes between wild-type and CRISPR-Cas9-mediated knockout cells. The candidates of interest are identified using bioinformatic analyses and verified by biochemical approaches. This protocol is highly versatile and applies to a variety of cells and different types of stimulation. For complete details on the use and execution of this protocol, please refer to (Zhu et al., 2021). Elsevier 2021-11-13 /pmc/articles/PMC8599494/ /pubmed/34820639 http://dx.doi.org/10.1016/j.xpro.2021.100962 Text en © 2021 The Author(s) 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
Zhu, Junji
Xiao, Wuhan
A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title_full A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title_fullStr A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title_full_unstemmed A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title_short A proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
title_sort proteomics protocol to identify stimulation-induced binding partners dependent on a specific gene in mammalian cells
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8599494/
https://www.ncbi.nlm.nih.gov/pubmed/34820639
http://dx.doi.org/10.1016/j.xpro.2021.100962
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