title: Sites and Functional Consequence of Alkylphenol Anesthetic Binding to Kv1.2 Channels.
abstract: Inhalational general anesthetics, such as sevoflurane and isoflurane, modulate a subset of brain Kv1 potassium channels. However, the Kv1.2 channel is resistant to propofol, a commonly used intravenous alkylphenol anesthetic. We hypothesize that propofol binds to a presumed pocket involving the channel's S4-S5 linker, but functional transduction is poor and, therefore, propofol efficacy is low. To test this hypothesis, we used a photoactive propofol analog (meta-aziPropofol = AziPm) to directly probe binding and electrophysiological and mutational analyses in Xenopus oocytes to probe function. We find that AziPm photolabels L321 in the S4-S5 linker of both the wild-type Kv1.2 and a mutant Kv1.2 (G329 T) with a novel gating phenotype. Furthermore, whereas propofol does not significantly modulate Kv1.2 WT but robustly potentiates Kv1.2 G329T, AziPm inhibits Kv1.2 WT and also potentiates Kv1.2 G329T. Kv1.2 modulation by AziPm was abolished by two mutations that decreased hydrophobicity at L321 (L321A and L321F), confirming the specific significance of the S4-S5 linker in the mechanism of general anesthetic modulation. Since AziPm binds to Kv1.2 G329T and shares the propofol ability to potentiate this mutant, the parent propofol likely also binds to the Kv1.2 channel. However, binding and alkylphenol-induced transduction are seemingly sensitive to the conformation of the S4-S5 linker site (altered by G329T) and subtle differences in the chemical structures of propofol and AziPm. Overall, the results are consistent with a mechanism of general anesthetic modulation that depends on the complementarity of necessary ligand binding and permissive ion channel conformations that dictate modulation and efficacy.
authors.affiliation: Department of Anesthesiology and Critical Care, University of Pennsylvania Perelman School of Medicine, 3620 Hamilton Walk, 305 John Morgan, Philadelphia, PA, 19104, USA.
authors.forename: Weiming
authors.initials: W
authors.lastname: Bu
authors.affiliation: Department of Neuroscience, Sidney Kimmel Medical College, Thomas Jefferson University, 900 Walnut Street, Room JHN 416, Philadelphia, PA, 19107, USA.
authors.forename: Qiansheng
authors.initials: Q
authors.lastname: Liang
authors.affiliation: Department of Neuroscience, Sidney Kimmel Medical College, Thomas Jefferson University, 900 Walnut Street, Room JHN 416, Philadelphia, PA, 19107, USA.
authors.forename: Lianteng
authors.initials: L
authors.lastname: Zhi
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, 19102, USA.
authors.forename: Lina
authors.initials: L
authors.lastname: Maciunas
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, 19102, USA.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Loll
authors.affiliation: Department of Anesthesiology and Critical Care, University of Pennsylvania Perelman School of Medicine, 3620 Hamilton Walk, 305 John Morgan, Philadelphia, PA, 19104, USA. roderic.eckenhoff@uphs.upenn.edu.
authors.forename: Roderic G
authors.initials: RG
authors.lastname: Eckenhoff
authors.affiliation: Department of Neuroscience, Sidney Kimmel Medical College, Thomas Jefferson University, 900 Walnut Street, Room JHN 416, Philadelphia, PA, 19107, USA. manuel.covarrubias@jefferson.edu.
authors.forename: Manuel
authors.initials: M
authors.lastname: Covarrubias
mesh_terms: D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D051663:Kv1.2 Potassium Channel / Q000378:metabolism*; D009865:Oocytes / Q000187:drug effects* / Q000378:metabolism; D015742:Propofol / Q000494:pharmacology*; D014981:Xenopus
chemical_list: D018685:Anesthetics, Inhalation; D051663:Kv1.2 Potassium Channel; D015742:Propofol
keywords: General anesthetic binding site; Kv channels; Propofol; S4–S5 linker