Affinity labeling of the protein kinase associated with the epidermal growth factor receptor in membrane vesicles from A431 cells.
Epidermal growth factor (EGF), a mitogenic polypeptide hormone, stimulates the phosphorylation of certain endogenous proteins in membrane preparations derived from A431 cells, a human tumor cell line. Membrane vesicles prepared from A431 cells were reacted with 5'-p-fluorosulfonylbenzoyl adenosine (5'-p-FSO2BzAdo). Reaction of the vesicles with 5'-p-FSO2BzAdo results in a time-dependent inhibition of EGF-stimulable protein kinase activity which parallels an increase in incorporation into the vesicles of the 5'-p-sulfonylbenzoyl-[8-14C]adenosine moiety from 5'-p-FSO2Bz[14C]Ado. The primary bands labeled have Mr = 170,000 and 150,000. Labeling of these bands by 5'-p-FSO2Bz[14C]Ado is inhibited by incubation of the membrane vesicles with adenyl-5'-yl imidodiphosphate, an ATP analog. Inactivation of the kinase with N-ethylmaleimide or by heating results in a sharply decreased labeling of the proteins with Mr = 170,000 and 150,000. Proteins of these molecular weights have previously been identified in these cells as the EGF receptor and a degradation product of the receptor. These experiments provide chemical evidence that the EGF receptor and the EGF-stimulable kinase are the same protein.
D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000345:Affinity Labels / Q000494:pharmacology*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000302:isolation & purification* / Q000494:pharmacology; D066246:ErbB Receptors; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D009369:Neoplasms / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000302:isolation & purification*; D011956:Receptors, Cell Surface / Q000302:isolation & purification*
D000345:Affinity Labels; D047428:Protein Kinase Inhibitors; D011956:Receptors, Cell Surface; D004815:Epidermal Growth Factor; C010959:5'-(4-fluorosulfonylbenzoyl)adenosine; D011494:Protein Kinases; D066246:ErbB Receptors; D000241:Adenosine