Binding, internalization, and intracellular processing of protein ligands. Derivation of rate constants by computer modeling.
Information about ligand binding, dissociation, internalization, and intracellular processing and about receptor turnover, processing, and insertion into the membrane is contained in the time-dependent changes in concentrations of membrane-associated and internalized ligand. Single experiments similar in design to those typically performed for Scatchard analyses of binding data conducted at physiological temperature and in the absence of inhibitors of ligand-receptor complex internalization and degradation can provide kinetic data sufficient to permit derivation of all the respective rate constants by numerical methods. We developed an analytical solution of the kinetic model which assumes that all of these processes follow first order kinetics. The model represents interactions of surface receptors (R)s, the surface ligand-receptor complex (LR)s and internalized receptor-ligand complex (LR)I: d[R]S/dt = Vr - kt[R]S - ka[L] [R]S + kd [LR]S; d[LR]S/dt = ka[L] [R]S - kd[LR]S - ke[LR]S; d[LR]I/dt = ke[LR]S - kh[LR]I; Vr is the constant rate of insertion of receptors into the membrane, kt is the internalization rate constant for free receptors, ka and kd are association and dissociation rate constants for ligand-surface receptor interaction, ke is the internalization rate constant for ligand-receptor complexes, and kh is the intracellular ligand decomposition rate constant. The interaction of radioiodinated human recombinant interferon-alpha 2a with the human alveolar lung carcinoma cell line, A549, was adequately accounted for by the model. The rate constants, numerically derived from time-dependent concentrations of surface-bound and internalized ligand of other systems taken from the literature, were in agreement with values of these rate constants individually measured by steady-state experiments. In cases where the fate of internalized radioactivity was more complex than assumed by the model, the parameters ka, kt, (kd + ke) and Vr could be derived from the time dependence of [LR]S.
D000230:Adenocarcinoma; D000818:Animals; D002460:Cell Line; D003198:Computer Simulation; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D007370:Interferon Type I / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008175:Lung Neoplasms; D008545:Melanoma; D008954:Models, Biological; D011506:Proteins / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism; D017471:Receptors, Interferon; D011994:Recombinant Proteins / Q000378:metabolism
D007370:Interferon Type I; D008024:Ligands; D011506:Proteins; D011971:Receptors, Immunologic; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D066246:ErbB Receptors