Kinetic model of the epidermal growth factor (EGF) receptor tyrosine kinase and a possible mechanism of its activation by EGF.
The tyrosine kinase activity of the epidermal growth factor receptor (EGFR-TK) was determined at varying poly-Glu6Ala3Tyr1 (GAT) or [Val5]-angiotensin II (AT) and constant ATP concentrations and vice versa. With GAT as substrate, double reciprocal plots intersected practically on the abscissa following EGFR-TK pre-activation with EGF, but below the abscissa without EGF pre-activation. The EGFR-TK inhibitors App(NH)p (5'-adenylyl-beta, gamma-imidodiphosphate) and ADP were competitive with ATP and noncompetitive with GAT. Four families of 1/v vs. 1/[ATP] plots, constructed at different fixed concentrations of ADP and a different constant concentration of GAT for each family, yielded Slope1/ATP replots which intersected to the left of the ordinate and below the abscissa. GAT and AT, as cosubstrates, were competitive with each other and noncompetitive with ATP; 1/v vs. 1/[GAT] or 1/[AT] plots were hyperbolic and reached horizontal asymptotes when v was expressed as the rate of common product formation. All data were subjected to computer best-fit analysis by a program written especially for this purpose. We conclude that (i) the EGFR-TK reaction follows a Sequential Bi-Bi Rapid Equilibrium Random mechanism, and (ii) EGF induces conformational changes in the EGFR-TK active center which lead to marked decreases in the apparent dissociation constants of both substrates of the kinase reaction and a concomitant increase in initial velocities and Vmax (apparent).
Posner
I
I
Department of Biological Chemistry, Hebrew University of Jerusalem, Israel.
Engel
M
M
Levitzki
A
A
D002294:Carcinoma, Squamous Cell; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008433:Mathematics; D008962:Models, Theoretical; D011505:Protein-Tyrosine Kinases / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases