The novel anticonvulsant drug, gabapentin (Neurontin), binds to the alpha(2)delta subunit of a calcium channel
The novel anticonvulsant drug, gabapentin (Neurontin), binds to the alpha(2)delta subunit of a calcium channel
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DOI：
10.1074/jbc.271.10.5768
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发表时间：
1996-03-08
影响因子：
4.8
通讯作者：
Woodruff, GN
中科院分区：
文献类型：
--
作者：
Gee, NS;Brown, JP;Woodruff, GN
加巴喷丁（1-（氨甲基）环己烷乙酸; Neurontin）是一种新型抗惊厥药物，其作用机制与其他抗癫痫药物明显不同。我们在这里报告的[H-3]加巴喷丁结合蛋白从猪大脑皮层膜的分离和表征。洗涤剂溶解的结合蛋白纯化1022倍，在六步柱层析程序，产率为3.9%。纯化的蛋白质具有130，000的表观亚基M（r），并且是高度糖基化的。M（r）130，000多肽EPFPSAVTIK的部分N-末端氨基酸序列与所报道的来自兔骨骼肌的L-型Ca 2+通道的α（2）δ亚基相同（汉密尔顿，S. L.，Hawkes，M. J.，Brush，K.，库克河，巴西-地昌河，巴西-地J.，Smilowitz，H. M.（1989）Biochemistry 28，7820-7828）。高水平的[H-3]加巴喷丁结合位点被发现在从大鼠脑，心脏和骨骼肌制备的膜。通过蛋白质印迹法测定，[H-3]加巴喷丁与转染α（2）δ cDNA的COS-7细胞的结合比对照组升高>10倍，这与α（2）δ蛋白的表达一致。最后，纯化的L-型Ca 2+通道复合物在解离条件下在离子交换柱上分级分离; [H-3]加巴喷丁结合活性密切跟随α（2）δ亚基的洗脱。[H-3]加巴喷丁是第一种与电压依赖性Ca 2+通道的α（2）δ亚基相互作用的药理学药物。
Gabapentin (1-(aminomethyl)cyclohexane acetic acid; Neurontin) is a novel anticonvulsant drug, with a mechanism of action apparently dissimilar to that of other antiepileptic agents. We report here the isolation and characterization of a [H-3]gabapentin binding protein from pig cerebral cortex membranes. The detergent-solubilized binding protein was purified 1022-fold, in a six-step column-chromatographic procedure, with a yield of 3.9%. The purified protein had an apparent subunit M(r) of 130,000, and was heavily glycosylated. The partial N-terminal amino acid sequence of the M(r) 130,000 polypeptide, EPFPSAVTIK, was identical to that reported for the alpha(2) delta subunit of the L-type Ca2+ channel from rabbit skeletal muscle (Hamilton, S. L., Hawkes, M. J., Brush, K., Cook, R., Chang, R. J., and Smilowitz, H. M. (1989) Biochemistry 28, 7820-7828). High levels of [H-3]gabapentin binding sites were found in membranes prepared from rat brain, heart and skeletal muscle. Binding of [H-3]gabapentin to COS-7 cells transfected with alpha(2) delta cDNA was elevated >10-fold over controls, consistent with the expression of alpha(2) delta protein, as measured by Western blotting. Finally, purified L-type Ca2+ channel complexes were fractionated, under dissociating conditions, on an ion-exchange column; [H-3]gabapentin binding activity closely followed the elution of the alpha(2) delta subunit. [H-3]Gabapentin is the first pharmacological agent described that interacts with an alpha(2) delta subunit of a voltage-dependent Ca2+ channel.