Phoneutria nigriventer toxin 1:: A novel, state-dependent inhibitor of neuronal sodium channels that interacts with μ conotoxin binding sites
Phoneutria nigriventer toxin 1:: A novel, state-dependent inhibitor of neuronal sodium channels that interacts with μ conotoxin binding sites
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DOI：
10.1124/mol.105.021147
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发表时间：
2006-06-01
影响因子：
3.6
通讯作者：
Van Renterghem, Catherine
中科院分区：
文献类型：
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作者：
Martin-Moutot, Nicole;Mansuelle, Pascal;Van Renterghem, Catherine
从南美武装蜘蛛Phoneutria nigriventer的毒液中纯化出一种毒素，其分子量为8600 Da，C端有酰胺化的甘氨酸残基。它似乎与之前从该毒液中分离出的毒素 1 (Tx1) 相同。 Tx1 可逆地抑制表达重组钠 (Na-v 1.2) 通道的中国仓鼠卵巢细胞中的钠电流，而不影响其快速生物物理特性。峰值钠电流的抑制动力学随膜电位的变化而变化，在-100至-50 mV范围内，随着去极化保持电位的增加，结合速率增加，解离速率降低。因此，随着膜去极化，Tx1 对通道的表观亲和力增加。单[I-125] iodo-Tx1 衍生物与大鼠脑膜中的一类位点（K-D = 80 pM，B-max = 0.43 pmol/mg 蛋白质）表现出高亲和力结合。通过针对钠通道 α 亚基中保守基序的抗体对溶解的结合位点进行免疫沉淀。 I-125-Tx1 结合被 mu 芋螺毒素 GIIIB 竞争性取代（IC50 = 0.5 mu M），但不被 1 mu M 河豚毒素取代。然而，在河豚毒素 (1 mu M) 存在的情况下，μ 芋螺毒素 GIIIB 对 I-125-Tx1 结合的抑制作用被消除。膜片钳和结合数据表明 P. nigriventer Tx1 是一种新型的状态依赖性钠通道阻滞剂，它与药理学位点 1 附近的位点结合，与 mu 芋螺毒素结合位点重叠，但与河豚毒素结合位点不重叠。
A toxin was purified to homogeneity from the venom of the South American armed spider Phoneutria nigriventer and found to have a molecular mass of 8600 Da and a C-terminally amidated glycine residue. It appears to be identical to Toxin 1 (Tx1) isolated previously from this venom. Tx1 reversibly inhibited sodium currents in Chinese hamster ovary cells expressing recombinant sodium (Na-v 1.2) channels without affecting their fast biophysical properties. The kinetics of inhibition of peak sodium current varied with membrane potential, with on-rates increasing and off-rates decreasing with more depolarized holding potentials in the -100 to -50 mV range. Thus, the apparent affinity of Tx1 for the channel increases as the membrane is depolarized. A mono[I-125] iodo-Tx1 derivative displayed high-affinity binding to a single class of sites (K-D = 80 pM, B-max = 0.43 pmol/mg protein) in rat brain membranes. Solubilized binding sites were immunoprecipitated by antibodies directed against a conserved motif in sodium channel alpha subunits. I-125-Tx1 binding was competitively displaced by mu conotoxin GIIIB (IC50 = 0.5 mu M) but not by 1 mu M tetrodotoxin. However, the inhibition of I-125-Tx1 binding by mu conotoxin GIIIB was abrogated in the presence of tetrodotoxin (1 mu M). Patch clamp and binding data indicate that P. nigriventer Tx1 is a novel, state-dependent sodium-channel blocker that binds to a site in proximity to pharmacological site 1, overlapping mu conotoxin but not tetrodotoxin binding sites.