title: Cloning, synthesis, and characterization of αO-conotoxin GeXIVA, a potent α9α10 nicotinic acetylcholine receptor antagonist.
abstract: We identified a previously unidentified conotoxin gene from Conus generalis whose precursor signal sequence has high similarity to the O1-gene conotoxin superfamily. The predicted mature peptide, αO-conotoxin GeXIVA (GeXIVA), has four Cys residues, and its three disulfide isomers were synthesized. Previously pharmacologically characterized O1-superfamily peptides, exemplified by the US Food and Drug Administration-approved pain medication, ziconotide, contain six Cys residues and are calcium, sodium, or potassium channel antagonists. However, GeXIVA did not inhibit calcium channels but antagonized nicotinic AChRs (nAChRs), most potently on the α9α10 nAChR subtype (IC50 = 4.6 nM). Toxin blockade was voltage-dependent, and kinetic analysis of toxin dissociation indicated that the binding site of GeXIVA does not overlap with the binding site of the competitive antagonist α-conotoxin RgIA. Surprisingly, the most active disulfide isomer of GeXIVA is the bead isomer, comprising, according to NMR analysis, two well-resolved but uncoupled disulfide-restrained loops. The ribbon isomer is almost as potent but has a more rigid structure built around a short 310-helix. In contrast to most α-conotoxins, the globular isomer is the least potent and has a flexible, multiconformational nature. GeXIVA reduced mechanical hyperalgesia in the rat chronic constriction injury model of neuropathic pain but had no effect on motor performance, warranting its further investigation as a possible therapeutic agent.
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China; luosulan2003@163.com.
authors.forename: Sulan
authors.initials: S
authors.lastname: Luo
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China;
authors.forename: Dongting
authors.initials: D
authors.lastname: Zhangsun
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia;
authors.forename: Peta J
authors.initials: PJ
authors.lastname: Harvey
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia;
authors.forename: Quentin
authors.initials: Q
authors.lastname: Kaas
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China;
authors.forename: Yong
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China;
authors.forename: Xiaopeng
authors.initials: X
authors.lastname: Zhu
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China;
authors.forename: Yuanyan
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou Hainan 570228, China;
authors.forename: Xiaodan
authors.initials: X
authors.lastname: Li
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow 117997, Russia;
authors.forename: Victor I
authors.initials: VI
authors.lastname: Tsetlin
authors.affiliation: Department of Biology and Psychiatry, University of Utah, Salt Lake City, UT 84112;
authors.forename: Sean
authors.initials: S
authors.lastname: Christensen
authors.affiliation: Department of Biology and Psychiatry, University of Utah, Salt Lake City, UT 84112;
authors.forename: Haylie K
authors.initials: HK
authors.lastname: Romero
authors.affiliation: Department of Biology and Psychiatry, University of Utah, Salt Lake City, UT 84112;
authors.forename: Melissa
authors.initials: M
authors.lastname: McIntyre
authors.affiliation: Department of Biology and Psychiatry, University of Utah, Salt Lake City, UT 84112;
authors.forename: Cheryl
authors.initials: C
authors.lastname: Dowell
authors.affiliation: Department of Pharmacology, Kirksville College of Osteopathic Medicine, A. T. Still University, Kirksville, MO 63501;
authors.forename: James C
authors.initials: JC
authors.lastname: Baxter
authors.affiliation: Department of Pharmacology, Kirksville College of Osteopathic Medicine, A. T. Still University, Kirksville, MO 63501;
authors.forename: Keith S
authors.initials: KS
authors.lastname: Elmslie
authors.affiliation: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia;
authors.forename: David J
authors.initials: DJ
authors.lastname: Craik
authors.affiliation: Department of Biology and Psychiatry, University of Utah, Salt Lake City, UT 84112; George E. Wahlen Veterans Affairs Medical Center, Salt Lake City, UT 84108; Department of Psychiatry, University of Utah, Salt Lake City, UT 84112.
authors.forename: J Michael
authors.initials: JM
authors.lastname: McIntosh
mesh_terms: D000577:Amides / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015220:Calcium Channels / Q000737:chemistry; D003001:Cloning, Molecular; D020916:Conotoxins / Q000737:chemistry*; D052078:Conus Snail / Q000737:chemistry*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006930:Hyperalgesia / Q000188:drug therapy; D020128:Inhibitory Concentration 50; D008297:Male; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009437:Neuralgia / Q000628:therapy; D018733:Nicotinic Antagonists / Q000737:chemistry*; D009865:Oocytes / Q000166:cytology; D011487:Protein Conformation; D021382:Protein Sorting Signals; D051381:Rats; D017207:Rats, Sprague-Dawley; D011978:Receptors, Nicotinic / Q000737:chemistry*; D014982:Xenopus laevis
chemical_list: D000577:Amides; D015220:Calcium Channels; C470699:Chrna10 protein, rat; C498541:Chrna9 protein, rat; D020916:Conotoxins; D018733:Nicotinic Antagonists; D021382:Protein Sorting Signals; D011978:Receptors, Nicotinic; C000602314:alpha-conotoxin, Conus generalis
keywords: NMR; nicotinic; pain; α9α10 nAChR; αO-conotoxin GeXIVA