title: ω-Tbo-IT1-New Inhibitor of Insect Calcium Channels Isolated from Spider Venom.
abstract: Novel disulfide-containing polypeptide toxin was discovered in the venom of the Tibellus oblongus spider. We report on isolation, spatial structure determination and electrophysiological characterization of this 41-residue toxin, called ω-Tbo-IT1. It has an insect-toxic effect with LD50 19 μg/g in experiments on house fly Musca domestica larvae and with LD50 20 μg/g on juvenile Gromphadorhina portentosa cockroaches. Electrophysiological experiments revealed a reversible inhibition of evoked excitatory postsynaptic currents in blow fly Calliphora vicina neuromuscular junctions, while parameters of spontaneous ones were not affected. The inhibition was concentration dependent, with IC50 value 40 ± 10 nM and Hill coefficient 3.4 ± 0.3. The toxin did not affect frog neuromuscular junctions or glutamatergic and GABAergic transmission in rat brains. Ca(2+) currents in Calliphora vicina muscle were not inhibited, whereas in Periplaneta americana cockroach neurons at least one type of voltage gated Ca(2+) current was inhibited by ω-Tbo-IT1. Thus, the toxin apparently acts as an inhibitor of presynaptic insect Ca(2+) channels. Spatial structure analysis of the recombinant ω-Tbo-IT1 by NMR spectroscopy in aqueous solution revealed that the toxin comprises the conventional ICK fold containing an extended β-hairpin loop and short β-hairpin loop which are capable of making "scissors-like mutual motions".
authors.affiliation: Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.
authors.forename: Alexander N
authors.initials: AN
authors.lastname: Mikov
authors.affiliation: Laboratory of Biophysics of Synaptic Processes, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Russia.
authors.forename: Irina M
authors.initials: IM
authors.lastname: Fedorova
authors.affiliation: Laboratory of Biophysics of Synaptic Processes, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Russia.
authors.forename: Natalia N
authors.initials: NN
authors.lastname: Potapieva
authors.affiliation: Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.
authors.forename: Ekaterina E
authors.initials: EE
authors.lastname: Maleeva
authors.affiliation: Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.
authors.forename: Yaroslav A
authors.initials: YA
authors.lastname: Andreev
authors.affiliation: Laboratory of Biophysics of Synaptic Processes, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Russia.
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Zaitsev
authors.affiliation: Laboratory of Biophysics of Synaptic Processes, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Russia.
authors.forename: Kira K
authors.initials: KK
authors.lastname: Kim
authors.affiliation: Department of Structural Biology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.|NBIC Centre, NRC "Kurchatov Institute", Russia.
authors.forename: Eduard V
authors.initials: EV
authors.lastname: Bocharov
authors.affiliation: NBIC Centre, NRC "Kurchatov Institute", Russia.
authors.forename: Timur N
authors.initials: TN
authors.lastname: Bozin
authors.affiliation: NBIC Centre, NRC "Kurchatov Institute", Russia.
authors.forename: Dmitry A
authors.initials: DA
authors.lastname: Altukhov
authors.affiliation: NBIC Centre, NRC "Kurchatov Institute", Russia.
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Lipkin
authors.affiliation: Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Kozlov
authors.affiliation: Laboratory of Biophysics of Synaptic Processes, I.M. Sechenov Institute of Evolutionary Physiology and Biochemistry RAS, Russia.
authors.forename: Denis B
authors.initials: DB
authors.lastname: Tikhonov
authors.affiliation: Department of Molecular Neurobiology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, Russia.
authors.forename: Eugene V
authors.initials: EV
authors.lastname: Grishin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001001:Anura; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000633:toxicity*; D015220:Calcium Channels / Q000737:chemistry / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003058:Cockroaches / Q000187:drug effects / Q000502:physiology; D004175:Diptera / Q000187:drug effects / Q000502:physiology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology; D015870:Gene Expression; D019476:Insect Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000633:toxicity*; D007814:Larva / Q000187:drug effects / Q000502:physiology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D016415:Sequence Alignment; D013111:Spider Venoms / Q000096:biosynthesis / Q000737:chemistry*; D013112:Spiders / Q000737:chemistry* / Q000502:physiology
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D019476:Insect Proteins; D011994:Recombinant Proteins; D013111:Spider Venoms; D002118:Calcium