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Identifying Protein Binding Partners of Phosphorylated Cortactin
Author(s) -
Johnson Jennifer,
Shaw Karen,
Kruchten Anne E.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1048.6
Subject(s) - cortactin , phosphorylation , microbiology and biotechnology , blot , protein phosphorylation , motility , binding protein , biology , chemistry , biochemistry , cytoskeleton , cell , gene , protein kinase a
Cortactin is an actin binding protein that is overexpressed and phosphorylated in several cancers resulting in altered rates of metastases, however no definitive role for cortactin phosphorylation has been determined. Cortactin interacts with several cellular binding partners, although only a few interactions thus far have been reported to be affected by protein phosphorylation. We hypothesize that the phosphorylation of cortactin results in different binding partners and cellular functions. Two approaches to determine the cellular binding partners of cortactin phosphorylation mutants were used. The first approach evaluated cortactin binding proteins by his‐tagged pull downs using Ni‐NTA beads, cellular extracts, and cortactin followed by separation on SDS‐PAGE and sequencing. The second approach utilized overlay assays and western blotting to identify the protein partners and observe the effects of the phosphorylation treatment. In the future, the differential binding proteins identified through both of these techniques will be tested to determine their effects on cellular motility. This work is supported by a Faculty Start‐Up award from the M.J. Murdock Charitable Trust to A. E. Kruchten.