Structure-function studies of the outer mouth and voltage sensor domain of hERG.

hERG has uniquely fast inactivation but slow activation processes. We study the structural basis for these unique gating properties using the following approaches: site-specific mutagenesis, the MTS accessibility test, disulfide bond formation, thermodynamic mutant cycle analysis, peptide toxin 'foot-printing', NMR spectroscopy, and molecular modelling. We propose the following: (1) two structural features in hERG's outer mouth contribute to its fast inactivation rate: a lack of 'open mouth'-stabilizing hydrogen bonds and an unusually long extracellular 'S5-P' linker that contains an alpha-helix. During membrane depolarization, four such 'S5-P helices' from the tetramer channel come near each other to occlude the outer mouth. This occurs rapidly due to the dynamic nature of the S5-P helices. (2) Two structural features in hERG's voltage sensor domain contribute to its slow activation rate: hERG's major voltage-sensor, S4, has three (instead of four as in Shaker) positive charges involved in gating charge transfer, and hERG has six (instead of three as in Shaker) negative charges in the other transmembrane segments (S1-S3) of the voltage sensor domain. Thus a less voltage-sensitive S4, in conjunction with more surrounding negative charges (some of which can form salt-bridges with S4's positive charges in the pre-open state), retards channel activation.