Apparent noncompetitive inhibition of choline transport in erythrocytes by inhibitors bound at the substrate site.

According to the conventional carrier model, an inhibitor bound at the substrate transfer site inhibits competitively when on the same side of the membrane as the substrate, but noncompetitively when on the opposite side. This prediction was tested with the nonpenetrating choline analog dimethyl-n-pentyl (2-hydroxyethyl) ammonium ion. In zero trans entry and infinite trans entry experiments, where the labeled substrate and the inhibitor occupy the same compartment, the inhibition was competitive, but in zero trans exit it was noncompetitive, in accord with the model. Similar behavior was seen with dimethyl-n-decyl (2-hydroxyethyl) ammonium ion. With this property of the choline transport system established, it becomes possible to estimate the relative affinity inside and outside of inhibitors present on both sides of the membrane. The tertiary amine, dibutylaminoethanol, which enters the cell by simple diffusion, is such an inhibitor. Here the inhibition kinetics were the reverse of those for nonpenetrating inhibitors; zero trans and infinite trans exit was inhibited competitively, and zero trans entry noncompetitively. It follows that dibutylaminoethanol binds predominantly to the inner carrier form.