TY - JOUR T1 - The Kinetics of Specific Glucocorticoid Binding in Rat Thymus Cytosol: Evidence for the Existence of Multiple Binding States JF - Molecular Pharmacology JO - Mol Pharmacol SP - 578 LP - 587 VL - 11 IS - 5 AU - JEFFREY L. KAINE AU - CARL J. NIELSEN AU - WILLIAM B. PRATT Y1 - 1975/09/01 UR - http://molpharm.aspetjournals.org/content/11/5/578.abstract N2 - The 27,000 x g supernatant prepared from rat thymocytes contains a steroid-binding protein that binds glucocorticoids in a specific, high-affinity manner. In this study the kinetics of binding of dexamethasone has been examined in order to provide an explanation for the very slow rate at which the binding equilibrium is achieved. The secondorder rate constant of association of 6 x 105 M-1 min-1 is much lower than that expected for a diffusion-limited process. Kinetic evidence is presented in support of the proposal that the apparent slow rate of binding is due to the initial rapid formation of a weak binding complex (dissociation constant = 1.75 x 10-7 M) followed by the production of a stronger complex. The first-order rate constant of formation for the second binding state is calculated to be 0.137 min-1, and the measured rate constant of dissociation is 4.6 x 10-3 min-1. On extended incubation a third, very tight binding state is produced. ACKNOWLEDGMENT The authors are indebted to Dr. Jules A. Shafer of the Department of Biological Chemistry, The University of Michigan, for his advice and criticism. ER -