Biotin identification proteomics in three-dimensional organotypic human skin cultures

Calvin J. Cable; Nihal Kaplan; Spiro Getsios; Paul M. Thomas; Bethany E. Perez White

doi:10.1007/7651_2019_239

Biotin identification proteomics in three-dimensional organotypic human skin cultures

Calvin J. Cable, Nihal Kaplan, Spiro Getsios, Paul M. Thomas, Bethany E. Perez White^*

^*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceeding › Chapter

1 Scopus citations

Abstract

Biotin identification (BioID) proteomics facilitates the unbiased detection of protein interaction neighborhoods in live cells. The BioID technique relies on the covalent biotin alteration of vicinal proteins by a modified bacterial biotin ligase. The biotin ligase is fused to a protein of interest to identify putative protein-protein interactions. Here, we describe the adaptation of this technique for use in three-dimensional epidermal cultures. Due to the covalent biotin modification of proteins, our protocol allows for the complete solubilization of the total cellular protein content in differentiated keratinocytes. Thus, a comprehensive network of potential interactors of a protein of interest can be mapped.

Original language	English (US)
Title of host publication	Methods in Molecular Biology
Publisher	Humana Press Inc.
Pages	185-197
Number of pages	13
DOIs	https://doi.org/10.1007/7651_2019_239
State	Published - 2020

Publication series

Name	Methods in Molecular Biology
Volume	2109
ISSN (Print)	1064-3745
ISSN (Electronic)	1940-6029

Funding

This work was supported by NIH grant AR072773, a Dermatology Foundation Career Development Award grant, and a Chicago Biomedical Consortium Postdoctoral Award to B.E.P.W. and NIH grant AR062110 to S.G. Work was performed with the support of the Northwestern University Skin Disease Research Center funded by NIH grant AR057216 and Northwestern Proteomics funded by NIH grants CA060553 and GM108569.

Keywords

3D skin culture
BioID
Biotin-identification proteomics
Epidermis
Keratinocytes
Protein-protein interaction

ASJC Scopus subject areas

Molecular Biology
Genetics

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10.1007/7651_2019_239

Cite this

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title = "Biotin identification proteomics in three-dimensional organotypic human skin cultures",

abstract = "Biotin identification (BioID) proteomics facilitates the unbiased detection of protein interaction neighborhoods in live cells. The BioID technique relies on the covalent biotin alteration of vicinal proteins by a modified bacterial biotin ligase. The biotin ligase is fused to a protein of interest to identify putative protein-protein interactions. Here, we describe the adaptation of this technique for use in three-dimensional epidermal cultures. Due to the covalent biotin modification of proteins, our protocol allows for the complete solubilization of the total cellular protein content in differentiated keratinocytes. Thus, a comprehensive network of potential interactors of a protein of interest can be mapped.",

keywords = "3D skin culture, BioID, Biotin-identification proteomics, Epidermis, Keratinocytes, Protein-protein interaction",

author = "Cable, {Calvin J.} and Nihal Kaplan and Spiro Getsios and Thomas, {Paul M.} and {Perez White}, {Bethany E.}",

note = "Funding Information: This work was supported by NIH grant AR072773, a Dermatology Foundation Career Development Award grant, and a Chicago Biomedical Consortium Postdoctoral Award to B.E.P.W. and NIH grant AR062110 to S.G. Work was performed with the support of the Northwestern University Skin Disease Research Center funded by NIH grant AR057216 and Northwestern Proteomics funded by NIH grants CA060553 and GM108569. Publisher Copyright: {\textcopyright} Springer Science+Business Media New York 2019.",

year = "2020",

doi = "10.1007/7651_2019_239",

language = "English (US)",

series = "Methods in Molecular Biology",

publisher = "Humana Press Inc.",

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T1 - Biotin identification proteomics in three-dimensional organotypic human skin cultures

AU - Cable, Calvin J.

AU - Kaplan, Nihal

AU - Getsios, Spiro

AU - Thomas, Paul M.

AU - Perez White, Bethany E.

N1 - Funding Information: This work was supported by NIH grant AR072773, a Dermatology Foundation Career Development Award grant, and a Chicago Biomedical Consortium Postdoctoral Award to B.E.P.W. and NIH grant AR062110 to S.G. Work was performed with the support of the Northwestern University Skin Disease Research Center funded by NIH grant AR057216 and Northwestern Proteomics funded by NIH grants CA060553 and GM108569. Publisher Copyright: © Springer Science+Business Media New York 2019.

PY - 2020

Y1 - 2020

N2 - Biotin identification (BioID) proteomics facilitates the unbiased detection of protein interaction neighborhoods in live cells. The BioID technique relies on the covalent biotin alteration of vicinal proteins by a modified bacterial biotin ligase. The biotin ligase is fused to a protein of interest to identify putative protein-protein interactions. Here, we describe the adaptation of this technique for use in three-dimensional epidermal cultures. Due to the covalent biotin modification of proteins, our protocol allows for the complete solubilization of the total cellular protein content in differentiated keratinocytes. Thus, a comprehensive network of potential interactors of a protein of interest can be mapped.

AB - Biotin identification (BioID) proteomics facilitates the unbiased detection of protein interaction neighborhoods in live cells. The BioID technique relies on the covalent biotin alteration of vicinal proteins by a modified bacterial biotin ligase. The biotin ligase is fused to a protein of interest to identify putative protein-protein interactions. Here, we describe the adaptation of this technique for use in three-dimensional epidermal cultures. Due to the covalent biotin modification of proteins, our protocol allows for the complete solubilization of the total cellular protein content in differentiated keratinocytes. Thus, a comprehensive network of potential interactors of a protein of interest can be mapped.

KW - 3D skin culture

KW - BioID

KW - Biotin-identification proteomics

KW - Epidermis

KW - Keratinocytes

KW - Protein-protein interaction

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T3 - Methods in Molecular Biology

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EP - 197

BT - Methods in Molecular Biology

PB - Humana Press Inc.

ER -

Biotin identification proteomics in three-dimensional organotypic human skin cultures

Abstract

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Keywords

ASJC Scopus subject areas

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