title: Amino-terminal determinants of U-type inactivation of voltage-gated K+ channels.
abstract: The T1 domain is a cytosolic NH2-terminal domain present in all Kv (voltage-dependent potassium) channels, and is highly conserved between Kv channel subfamilies. Our characterization of a truncated form of Kv1.5 (Kv1.5deltaN209) expressed in myocardium demonstrated that deletion of the NH2 terminus of Kv1.5 imparts a U-shaped inactivation-voltage relationship to the channel, and prompted us to investigate the NH2 terminus as a regulatory site for slow inactivation of Kv channels. We examined the macroscopic inactivation properties of several NH2-terminal deletion mutants of Kv1.5 expressed in HEK 293 cells, demonstrating that deletion of residues up to the T1 boundary (Kv1.5deltaN19, Kv1.5deltaN91, and Kv1.5deltaN119) did not alter Kv1.5 inactivation, however, deletion mutants that disrupted the T1 structure consistently exhibited inactivation phenotypes resembling Kv1.5deltaN209. Chimeric constructs between Kv1.5 and the NH2 termini of Kv1.1 and Kv1.3 preserved the inactivation kinetics observed in full-length Kv1.5, again suggesting that the Kv1 T1 domain influences slow inactivation. Furthermore, disruption of intersubunit T1 contacts by mutation of residues Glu(131) and Thr(132) to alanines resulted in channels exhibiting a U-shaped inactivation-voltage relationship. Fusion of the NH2 terminus of Kv2.1 to the transmembrane segments of Kv1.5 imparted a U-shaped inactivation-voltage relationship to Kv1.5, whereas fusion of the NH2 terminus of Kv1.5 to the transmembrane core of Kv2.1 decelerated Kv2.1 inactivation and abolished the U-shaped voltage dependence of inactivation normally observed in Kv2.1. These data suggest that intersubunit T1 domain interactions influence U-type inactivation in Kv1 channels, and suggest a generalized influence of the T1 domain on U-type inactivation between Kv channel subfamilies.
authors.affiliation: Department of Physiology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
authors.forename: Harley T
authors.initials: HT
authors.lastname: Kurata
authors.forename: Gordon S
authors.initials: GS
authors.lastname: Soon
authors.forename: Jodene R
authors.initials: JR
authors.lastname: Eldstrom
authors.forename: Grace W K
authors.initials: GW
authors.lastname: Lu
authors.forename: David F
authors.initials: DF
authors.lastname: Steele
authors.forename: David
authors.initials: D
authors.lastname: Fedida
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D051668:Delayed Rectifier Potassium Channels; D004594:Electrophysiology; D017353:Gene Deletion; D006801:Humans; D007700:Kinetics; D051666:Kv1.5 Potassium Channel; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010641:Phenotype; D017354:Point Mutation; D011188:Potassium / Q000737:chemistry; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D024642:Potassium Channels, Voltage-Gated; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D051669:Shab Potassium Channels; D014162:Transfection
chemical_list: D051668:Delayed Rectifier Potassium Channels; C496981:KCNA5 protein, human; C496988:KCNB1 protein, human; D051666:Kv1.5 Potassium Channel; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011993:Recombinant Fusion Proteins; D051669:Shab Potassium Channels; D011188:Potassium