The Journal of pharmacology and experimental therapeutics

Different state dependencies of 4-aminopyridine binding to rKv1.4 and rKv4.2: role of the cytoplasmic halves of the fifth and sixth transmembrane segments.

PMID 10411564


4-Aminopyridine (4AP) binding to rKv1.4 occurs preferentially in the activated state, whereas binding to rKv4.2 occurs in the rested state. To explore structural basis for the different state dependencies of 4AP binding, regions of rKv1.4 that are likely to form the 4AP-binding site and/or the activation gate were replaced by the corresponding rKv4.2 sequences one at a time, and the resulting effects on channel gating and 4AP binding were examined. Replacing the amino acid sequence of rKv1.4 in the intracellular loop between the fourth and fifth transmembrane segments (S4 and S5) with that of rKv4.2 did not alter channels' gating properties or the state dependence of 4AP binding. On the other hand, replacing the rKv1.4 sequence in the cytoplasmic half of S5 (N-S5) or S6 (C-S6) with that of rKv4.2 markedly altered the voltage dependence and kinetics of activation gate function. Importantly, these mutations transferred the rested-state 4AP-binding preference from the donor to the host channel. These data can be explained by a scheme in which the function of the activation gate determines the state dependence of 4AP binding, although the relationship between the binding site and the gate may be similar between rKv1.4 and rKv4.2. The amino acid sequences in the N-S5 and C-S6 domains contribute to this activation gate function.