Kinetic properties of Ash/Grb2-interacting GDP/GTP exchange protein

Ash/Grb2 is a protein having one SH2 domain flanked by two SH3 domains and is implicated to serve as an adaptor protein which links the EGF receptor to mammalian Sos (mSos), a GDP/GTP exchange protein for Ras. We isolated here several Ash-interacting proteins from bovine brain cytosol by use of glutathione-S-transferase-Ash-linked agarose column chromatography. The Ash-interacting proteins stimulated the GDP/GTP exchange reaction of Ki-Ras and Ha-Ras but not that of other small GTP-binding proteins including at least Rap1, RhoA, Rac1, and Rab3A. The Ash-interacting proteins were much more active on the post-translationally lipid-modified form of Ki-Ras than on the unmodified form. At least one of them was identified as mSos by Western blot analysis using a specific anti-mSos antibody.