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Mapping the site of interaction between annexin VI and the p120 GAP C2 domain
Author(s) -
Chow Andrew,
Gawler Debra
Publication year - 1999
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)01336-8
Subject(s) - annexin a2 , annexin , linker , immunoprecipitation , binding site , fusion protein , c2 domain , chemistry , protein–protein interaction , binding protein , microbiology and biotechnology , biology , biochemistry , cell , membrane , operating system , computer science , gene , recombinant dna
Annexin VI is a Ca 2+ ‐dependent membrane and phospholipid binding protein. It mediates a protein‐protein interaction with the Ras p21 regulatory protein p120 GAP . In this study we have mapped the binding site of GAP within the annexin VI protein. Using Far Western overlay binding assays and cell lysate competition studies we have mapped the site of interaction to the inter‐lobe linker region; amino acids 325–363. Finally, using a GST fusion protein corresponding to this linker region we have demonstrated that cellular loading of the fusion protein into Rat‐1 fibroblasts by electroporation blocks the interaction and co‐immunoprecipitation of annexin VI and GAP.