title: Calmodulin inhibits the epidermal growth factor receptor tyrosine kinase.
abstract: We demonstrate in this report that the epidermal growth factor (EGF) receptor from rat liver can be isolated by calmodulin affinity chromatography by binding in the presence of Ca2+ and elution with a Ca(2+)-chelating agent. The bulk of the EGF receptor is not eluted by a NaCl gradient in the presence of Ca2+. We ascertained the identity of the isolated receptor by immunoblot and immunoprecipitation using a polyclonal antibody against an EGF receptor from human origin. The purified receptor is autophosphorylated in tyrosine residues in an EGF-stimulated manner, and EGF-dependent phosphorylation of serine residues was also detected. Both the EGF and the transforming growth factor-alpha stimulate the tyrosine-directed protein kinase activity of the isolated receptor with similar affinities. Furthermore, we demonstrate that calmodulin inhibits the EGF-dependent tyrosine-directed protein kinase activity associated to the receptor in a concentration-dependent manner. This inhibition is partially Ca2+ dependent and is not displaced by increasing the concentration of EGF up to an EGF/calmodulin ratio of 10 (mol/mol). In addition, calmodulin was phosphorylated in an EGF-stimulated manner in the presence of a basic protein (histone) as cofactor and in the absence, but not in the presence, of Ca2+.
authors.affiliation: Instituto de Investigaciones Biomédicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
authors.forename: E
authors.initials: E
authors.lastname: San José
authors.forename: A
authors.initials: A
authors.lastname: Benguría
authors.forename: P
authors.initials: P
authors.lastname: Geller
authors.forename: A
authors.initials: A
authors.lastname: Villalobo
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002147:Calmodulin / Q000494:pharmacology*; D002462:Cell Membrane / Q000201:enzymology / Q000378:metabolism; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D066246:ErbB Receptors; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D008297:Male; D010766:Phosphorylation; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D051381:Rats; D011919:Rats, Inbred Strains; D016211:Transforming Growth Factor alpha / Q000494:pharmacology
chemical_list: D002147:Calmodulin; D016211:Transforming Growth Factor alpha; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases