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Curr Protoc Protein Sci. 2013 Apr;Chapter 27:Unit 27.4. doi: 10.1002/0471140864.ps2704s72.

Preparation of recombinant protein spotted arrays for proteome-wide identification of kinase targets.

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  • 1Department of Genetics, Stanford University, Stanford, California, USA.

Abstract

Protein microarrays allow unique approaches for interrogating global protein interaction networks. Protein arrays can be divided into two categories: antibody arrays and functional protein arrays. Antibody arrays consist of various antibodies and are appropriate for profiling protein abundance and modifications. Functional full-length protein arrays employ full-length proteins with various post-translational modifications. A key advantage of the latter is rapid parallel processing of large number of proteins for studying highly controlled biochemical activities, protein-protein interactions, protein-nucleic acid interactions, and protein-small molecule interactions. This unit presents a protocol for constructing functional yeast protein microarrays for global kinase substrate identification. This approach enables the rapid determination of protein interaction networks in yeast on a proteome-wide level. The same methodology can be readily applied to higher eukaryotic systems with careful consideration of overexpression strategy.

PMID:
23546622
PMCID:
PMC4414062
DOI:
10.1002/0471140864.ps2704s72
[PubMed - indexed for MEDLINE]
Free PMC Article
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