Figure 8

Homologous competition of [³H]EB at the low affinity site is substantially different from competition at the high affinity site when ligand depletion is significant. A and B. Homologous competition from the high affinity site (A) and the low affinity site (B) with [³H]EB = 0.013 nM and R1T = 0.00013 nM leads to negligible depletion of [³H]EB. The competition curves are sigmoidal. C and D. [³H]EB = 0.013 nM and R1T = 130 nM lead to significant ligand depletion. Competition at the high affinity site (C) with ligand depletion is a distorted sigmoid curve similar to the total competition curves at high ligand depletion in Figure 7. In contrast, competition at the low affinity site (D) is a peak with maximum binding at 190 nM cold EB. From the right-hand scales of A and B with the right-hand scales of C and D, ligand depletion changes the fractional contribution of the low affinity site to the total binding. The low affinity site contributes less than 3.5 × 10^-4 of the maximum total binding when ligand depletion is negligible (A and B). In contrast, the low affinity site contributes more than 0.08 of the maximum total binding with significant ligand depletion (C and D).