title: μ-conotoxin KIIIA derivatives with divergent affinities versus efficacies in blocking voltage-gated sodium channels.
abstract: The possibility of independently manipulating the affinity and efficacy of pore-blocking ligands of sodium channels is of interest for the development of new drugs for the treatment of pain. The analgesic mu-conotoxin KIIIA (KIIIA), a 16-residue peptide with three disulfide bridges, is a pore blocker of voltage-gated sodium channels, including neuronal subtype Na(V)1.2 (K(d) = 5 nM). At saturating concentrations, KIIIA incompletely blocks the sodium current of Na(V)1.2, leaving a 5% residual current (rI(Na)). Lys7 is an important residue: the K7A mutation decreases both the efficacy (i.e., increases rI(Na) to 23%) and the affinity of the peptide (K(d) = 115 nM). In this report, various replacements of residue 7 were examined to determine whether affinity and efficacy were inexorably linked. Because of their facile chemical synthesis, KIIIA analogues that had as a core structure the disulfide-depleted KIIIA[C1A,C2U,C9A,C15U] (where U is selenocysteine) or ddKIIIA were used. Analogues ddKIIIA and ddKIIIA[K7X], where X represents one of nine different amino acids, were tested on voltage-clamped Xenopus oocytes expressing rat Na(V)1.2 or Na(V)1.4. Their affinities ranged from 0.01 to 36 muM and rI(Na) values from 2 to 42%, and these two variables appeared to be uncorrelated. Instead, rI(Na) varied inversely with side chain size, and remarkably charge and hydrophobicity appeared to be inconsequential. The ability to manipulate a mu-conopeptide's affinity and efficacy, as well as its capacity to interfere with subsequent tetrodotoxin binding, greatly expands its scope as a reagent for probing sodium channel structure and function and may also lead to the development of mu-conotoxins as safe analgesics.
authors.affiliation: Department of Biology, University of Utah, Salt Lake City, Utah 84112, USA.
authors.forename: Min-Min
authors.initials: MM
authors.lastname: Zhang
authors.forename: Tiffany S
authors.initials: TS
authors.lastname: Han
authors.forename: Baldomero M
authors.initials: BM
authors.lastname: Olivera
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Bulaj
authors.forename: Doju
authors.initials: D
authors.lastname: Yoshikami
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D020916:Conotoxins / Q000138:chemical synthesis / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000502:physiology*; D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; D009865:Oocytes / Q000378:metabolism; D011485:Protein Binding; D011522:Protons; D051381:Rats; D017279:Selenocysteine / Q000235:genetics; D026941:Sodium Channel Blockers / Q000138:chemical synthesis / Q000378:metabolism*; D015222:Sodium Channels / Q000502:physiology*; D014981:Xenopus
chemical_list: D020916:Conotoxins; D004220:Disulfides; D007202:Indicators and Reagents; D009124:Muscle Proteins; D062551:NAV1.2 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; D011522:Protons; C568253:Scn2A protein, rat; C466180:Scn4a protein, rat; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D017279:Selenocysteine
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