文摘
Di-substituted cyclohexyl (DSC) derivatives inhibit the voltage-gated potassium channel, Kv1.3,and have immunosuppressant activity (Schmalhofer et al. (2002) Biochemistry 41, 7781-7794). Thisclass of inhibitors displays Hill coefficients of near 2 in functional assays, and trans DSC analoguesappear to selectively interact with Kv1.3 channel conformations related to C-type inactivation. To furtherunderstand the details of the DSC inhibitor interaction with potassium channels, trans-1-(N-n-propylcarbamoyloxy)-4-phenyl-4-(3-(2-methoxyphenyl)-3-oxo-2-azaprop-1-yl)cyclo-hexane (trans-NPCO-DSC) was radiolabeled with tritium, and its binding characteristics to Kv1.3 channels were determined.Specific binding of [3H]-trans-NPCO-DSC to Kv1.3 channels is a saturable, time-dependent, and fullyreversible process. Saturation binding isotherms and competition binding experiments are consistent withthe presence of two receptor sites for DSC derivatives on the Kv1.3 channel that display positive allostericcooperativity. The high affinity interaction of [3H]-trans-NPCO-DSC with Kv1.3 channels appears tocorrelate with the rates of C-type inactivation of the channel. These data, taken together, mark the firstdemonstration of the existence of multiple binding sites for an inhibitor of an ion channel and suggestthat the high affinity interaction of trans-NPCO-DSC and similar inhibitors with Kv1.3 channels could beexploited for the development of selective molecules that target this protein.