Dynamic coupling of voltage sensor and gate involved in closed-state inactivation of kv4.2 channels.
Voltage-gated potassium channels related to the Shal gene of Drosophila (Kv4 channels) mediate a subthreshold-activating current (I(SA)) that controls dendritic excitation and the backpropagation of action potentials in neurons. Kv4 channels also exhibit a prominent low voltage-induced closed-state inactivation, but the underlying molecular mechanism is poorly understood. Here, we examined a structural model in which dynamic coupling between the voltage sensors and the cytoplasmic gate underlies inactivation in Kv4.2 channels. We performed an alanine-scanning mutagenesis in the S4-S5 linker, the initial part of S5, and the distal part of S6 and functionally characterized the mutants under two-electrode voltage clamp in Xenopus oocytes. In a large fraction of the mutants (>80%) normal channel function was preserved, but the mutations influenced the likelihood of the channel to enter the closed-inactivated state. Depending on the site of mutation, low-voltage inactivation kinetics were slowed or accelerated, and the voltage dependence of steady-state inactivation was shifted positive or negative. Still, in some mutants these inactivation parameters remained unaffected. Double mutant cycle analysis based on kinetic and steady-state parameters of low-voltage inactivation revealed that residues known to be critical for voltage-dependent gate opening, including Glu 323 and Val 404, are also critical for Kv4.2 closed-state inactivation. Selective redox modulation of corresponding double-cysteine mutants supported the idea that these residues are involved in a dynamic coupling, which mediates both transient activation and closed-state inactivation in Kv4.2 channels.
Barghaan
Jan
J
Zentrum für Experimentelle Medizin, Institut für Vegetative Physiologie und Pathophysiologie, Universit ä tsklinikum Hamburg-Eppendorf, Hamburg, Germany.
Bähring
Robert
R
D019943:Amino Acid Substitution; D000818:Animals; D005260:Female; D006801:Humans; D015640:Ion Channel Gating / Q000235:genetics; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D008958:Models, Molecular; D009865:Oocytes; D018408:Patch-Clamp Techniques; D054730:Protein Interaction Domains and Motifs / Q000502:physiology*; D051672:Shal Potassium Channels / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D055672:Static Electricity; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D014982:Xenopus laevis
D051672:Shal Potassium Channels