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) |
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Title of host publication | Methods in Molecular Biology |
Publisher | Humana Press Inc. |
Pages | 185-197 |
Number of pages | 13 |
DOIs | |
State | Published - 2020 |
Publication series
Name | Methods in Molecular Biology |
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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