title: Interactions between the C-terminus of Kv1.5 and Kvβ regulate pyridine nucleotide-dependent changes in channel gating.
abstract: Voltage-gated potassium (Kv) channels are tetrameric assemblies of transmembrane Kv proteins with cytosolic N- and C-termini. The N-terminal domain of Kv1 proteins binds to β-subunits, but the role of the C-terminus is less clear. Therefore, we studied the role of the C-terminus in regulating Kv1.5 channel and its interactions with Kvβ-subunits. When expressed in COS-7 cells, deletion of the C-terminal domain of Kv1.5 did not affect channel gating or kinetics. Coexpression of Kv1.5 with Kvβ3 increased current inactivation, whereas Kvβ2 caused a hyperpolarizing shift in the voltage dependence of current activation. Inclusion of NADPH in the patch pipette solution accelerated the inactivation of Kv1.5-Kvβ3 currents. In contrast, NADP(+) decreased the rate and the extent of Kvβ3-induced inactivation and reversed the hyperpolarizing shift in the voltage dependence of activation induced by Kvβ2. Currents generated by Kv1.5ΔC+Kvβ3 or Kv1.5ΔC+Kvβ2 complexes did not respond to changes in intracellular pyridine nucleotide concentration, indicating that the C-terminus is required for pyridine nucleotide-dependent interactions between Kvβ and Kv1.5. A glutathione-S-transferase (GST) fusion protein containing the C-terminal peptide of Kv1.5 did not bind to apoKvβ2, but displayed higher affinity for Kvβ2:NADPH than Kvβ2:NADP(+). The GST fusion protein also precipitated Kvβ proteins from mouse brain lysates. Pull-down experiments, structural analysis and electrophysiological data indicated that a specific region of the C-terminus (Arg543-Val583) is required for Kvβ binding. These results suggest that the C-terminal domain of Kv1.5 interacts with β-subunits and that this interaction is essential for the differential regulation of Kv currents by oxidized and reduced nucleotides.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL 33612, USA. stippara@health.usf.edu
authors.forename: Srinivas M
authors.initials: SM
authors.lastname: Tipparaju
authors.forename: Xiao-Ping
authors.initials: XP
authors.lastname: Li
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Kilfoil
authors.forename: Bin
authors.initials: B
authors.lastname: Xue
authors.forename: Vladimir N
authors.initials: VN
authors.lastname: Uversky
authors.forename: Aruni
authors.initials: A
authors.lastname: Bhatnagar
authors.forename: Oleg A
authors.initials: OA
authors.lastname: Barski
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D017353:Gene Deletion; D015640:Ion Channel Gating / Q000502:physiology*; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D051666:Kv1.5 Potassium Channel / Q000235:genetics / Q000502:physiology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation / Q000235:genetics; D018408:Patch-Clamp Techniques; D021122:Protein Subunits / Q000502:physiology*; D011725:Pyridines / Q000378:metabolism*
chemical_list: D051666:Kv1.5 Potassium Channel; D021122:Protein Subunits; D011725:Pyridines; C023666:pyridine