title: Design and characterization of a highly selective peptide inhibitor of the small conductance calcium-activated K+ channel, SkCa2.
abstract: Apamin-sensitive small conductance calcium-activated potassium channels (SKCa1-3) mediate the slow afterhyperpolarization in neurons, but the molecular identity of the channel has not been defined because of the lack of specific inhibitors. Here we describe the structure-based design of a selective inhibitor of SKCa2. Leiurotoxin I (Lei) and PO5, peptide toxins that share the RXCQ motif, potently blocked human SKCa2 and SKCa3 but not SKCa1, whereas maurotoxin, Pi1, Tskappa, and PO1 were ineffective. Lei blocked these channels more potently than PO5 because of the presence of Ala(1), Phe(2), and Met(7). By replacing Met(7) in the RXCQ motif of Lei with the shorter, unnatural, positively charged diaminobutanoic acid (Dab), we generated Lei-Dab(7), a selective SKCa2 inhibitor (K(d) = 3.8 nm) that interacts with residues in the external vestibule of the channel. SKCa3 was rendered sensitive to Lei-Dab(7) by replacing His(521) with the corresponding SKCa2 residue (Asn(367)). Intracerebroventricular injection of Lei-Dab(7) into mice resulted in no gross central nervous system toxicity at concentrations that specifically blocked SKCa2 homotetramers. Lei-Dab(7) will be a useful tool to investigate the functional role of SKCa2 in mammalian tissues.
authors.affiliation: Department of Physiology and Biophysics, University of California, Irvine, California 92697, USA. vshakkot@uci.edu
authors.forename: V G
authors.initials: VG
authors.lastname: Shakkottai
authors.forename: I
authors.initials: I
authors.lastname: Regaya
authors.forename: H
authors.initials: H
authors.lastname: Wulff
authors.forename: Z
authors.initials: Z
authors.lastname: Fajloun
authors.forename: H
authors.initials: H
authors.lastname: Tomita
authors.forename: M
authors.initials: M
authors.lastname: Fathallah
authors.forename: M D
authors.initials: MD
authors.lastname: Cahalan
authors.forename: J J
authors.initials: JJ
authors.lastname: Gargus
authors.forename: J M
authors.initials: JM
authors.lastname: Sabatier
authors.forename: K G
authors.initials: KG
authors.lastname: Chandy
mesh_terms: D000409:Alanine / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine / Q000737:chemistry; D019556:COS Cells; D002460:Cell Line; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D006801:Humans; D007700:Kinetics; D008715:Methionine / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009498:Neurotoxins / Q000494:pharmacology; D016716:PC12 Cells; D010455:Peptides / Q000737:chemistry* / Q000494:pharmacology; D010649:Phenylalanine / Q000494:pharmacology; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000737:chemistry*; D024681:Potassium Channels, Calcium-Activated / Q000037:antagonists & inhibitors*; D011485:Protein Binding; D051381:Rats; D012604:Scorpion Venoms / Q000737:chemistry / Q000494:pharmacology*; D017386:Sequence Homology, Amino Acid; D051640:Small-Conductance Calcium-Activated Potassium Channels; D013912:Threonine / Q000737:chemistry; D014162:Transfection; D014633:Valine / Q000737:chemistry; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
chemical_list: D009498:Neurotoxins; D010455:Peptides; C104751:Pi1 toxin; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024681:Potassium Channels, Calcium-Activated; D012604:Scorpion Venoms; D051640:Small-Conductance Calcium-Activated Potassium Channels; C442227:diaminobutanoyl(7)-leiurotoxin I; C104498:maurotoxin; C080397:scorpion venom P05; D013912:Threonine; D010649:Phenylalanine; D005680:gamma-Aminobutyric Acid; D001120:Arginine; D008715:Methionine; D014633:Valine; D000409:Alanine