Abstract

Quinidine and 4AP are two nonspecific K channel blockers. Both block voltage-gated K channels from the intracellular side of the membrane and, in most cases, binding is facilitated by channel activation. However, there are distinct differences between quinidine and 4AP in the time- and voltage-dependencies of drug-channel interaction. To learn about the molecular basis underlying the similarities as well as differences in drug actions between quinidine and 4AP, we used rKv1.4 (rat isoform of Kv1.4) as a model and studied: 1) Is there an overlap between the binding sites of quinidine and 4AP? and 2) What factors are involved in determining the binding affinity and kinetics of drug-channel interaction? Our data show that mutations at a position in the S6 domain of rKv1.4 (position 529) can cause dramatic and often opposite effects on quinidine and 4AP binding. For quinidine, the degree of steric hindrance imposed by side chain at position 529 is an important factor in determining binding affinity. For 4AP, 529 mutations that slow the rate of deactivation reduce binding affinity, probably due to a low binding affinity in the open state. This, in conjunction with the observations that 4AP binding is facilitated by channel activation, suggests that optimal 4AP binding may occur in a transitional state between fully-closed and fully-open states. In addition, hydrophobic interactions between blocker molecules and residues at 529 tend to stabilize the binding of both quinidine and 4AP. Because the S6 amino acid sequences are well conserved among many voltage-gated K channels, our findings have general implications in understanding the structural determinants of quinidine and 4AP binding to different K channels.