Alteration of the kinetic properties of the epidermal growth factor receptor tyrosine kinase by basic proteins.
Previous studies have shown that lysine- and arginine-rich proteins can enhance the activity of tyrosine and serine/threonine protein kinases. However, the kinetics and mechanism of this activation are not fully understood. Therefore we investigated the ability of poly(amino acids) and the arginine-rich protein, protamine, to alter the kinetic properties of epidermal growth factor (EGF) receptor protein-tyrosine kinase activity using immunoaffinity-purified receptor isolated from human epidermoid carcinoma (A431) cells. Poly(L-lysine), poly(L-arginine) and protamine stimulated EGF receptor kinase activity by 3-5-fold at non-saturating doses of ATP and peptide substrate, while poly(L-glutamate) had no effect. Initial kinetic studies demonstrated an increase in the maximum velocity and a decrease in the apparent Km for the peptide substrate angiotensin II in the presence of the basic effectors. Further analysis of the kinetic mechanism by product inhibition revealed that protamine altered the pattern of ADP inhibition towards the peptide substrate but not towards ATP. The change was indicative of the receptor's ability to form an enzyme-angiotensin II-ADP ternary complex in the presence of protamine but not in its absence. In addition, the basic effectors had a substantially decreased influence on the kinase activity of a C-terminally truncated form of the EGF receptor. Thus the changes in kinase activity may be partially mediated by the C-terminal region of the receptor, which contains the sites of receptor self-phosphorylation. These results suggest that the basic domains of proteins can interact with the EGF receptor to induce changes in its kinetic properties, especially with regard to reactant recognition and binding.
Department of Physiological Chemistry, University of Wisconsin-Madison 53706.
L
L
Hubler
P S
PS
Leventhal
P J
PJ
Bertics
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D066246:ErbB Receptors / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000494:pharmacology*; D011099:Polyglutamic Acid / Q000494:pharmacology; D011107:Polylysine / Q000494:pharmacology; D011479:Protamines / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D010455:Peptides; D011479:Protamines; D000804:Angiotensin II; D011107:Polylysine; C015462:polyarginine; D011099:Polyglutamic Acid; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases