CONSTRAINING LIGAND-BINDING SITE STOICHIOMETRY SUGGESTS THAT A CYCLICNUCLEOTIDE-GATED CHANNEL IS COMPOSED OF 2 FUNCTIONAL DIMERS

Cyclic nucleotide-gated ion channels are composed of four pore-forming subunits. Binding of cyclic nucleotide to a site in the intracellular carboxyl terminus of each subunit leads to channel activation. Since there are four subunits, four binding events are possible. In this stu dy, we investigate the effects of individual binding events on activat ion by studying channels containing one, two, three, or four functiona l binding sites. The binding of a single ligand significantly increase s opening, although four ligands are required for full activation. The data are inconsistent with models in which the four subunits activate in a single concerted step (Monod-Wyman-Changeux model) or in four in dependent steps (Hodgkin-Huxley model). Instead, the four subunits may associate and activate as two independent dimers.