The sodium channel from rat brain. Purification and subunit composition.
The sodium channel from rat brain. Purification and subunit composition.
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DOI：
10.1016/s0021-9258(17)43460-0
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发表时间：
1984-02
期刊：
影响因子：
--
通讯作者：
R. Hartshorne;W. Catterall
中科院分区：
文献类型：
--
作者：
R. Hartshorne;W. Catterall
描述了一种从大鼠脑中纯化钠通道1380倍至基本均一的方法。将通道溶解在Triton X-100中，并通过加入磷脂酰胆碱和10 mM CaCl 2来稳定。通过在DEAE-Sephadex、羟基磷灰石和麦胚凝集素/Sepharose上连续层析，然后通过蔗糖梯度沉降来纯化。最终制剂结合2910 pmol石房蛤毒素（STX）/mg蛋白质或0.9 mol STX/mol Mr约316，000的钠通道。三个多肽亚基包含纯蛋白在十二烷基硫酸钠聚丙烯酰胺凝胶上银染色强度的90%，并在蔗糖梯度沉降中作为与STX结合活性一致的化学计量复合物迁移：α与Mr约260，000，β 1与Mr约39，000，β 2与Mr约37，000。α亚基，纯化和完整的突触体中，表现出异常高的外推电泳自由流动性的十二烷基硫酸钠-聚丙烯酰胺凝胶电泳过程中表现出不良的行为。提出了α 1（β 1）1（β 2）1的亚基化学计量。
A procedure is described for purification of the sodium channel 1380-fold from rat brain to essential homogeneity. The channel is solubilized in Triton X-100 and stabilized by addition of phosphatidylcholine and 10 mM CaCl2. It is purified by sequential chromatography on DEAE-Sephadex, hydroxylapatite, and wheat germ agglutinin/Sepharose followed by sedimentation through sucrose gradients. The final preparation binds 2910 pmol of saxitoxin (STX)/mg of protein or 0.9 mol of STX/mol of sodium channel of Mr approximately 316,000. Three polypeptide subunits comprise 90% of the silver stain intensity on sodium dodecyl sulfatepolyacrylamide gels of the pure protein and migrate as a stoichiometric complex coincident with STX-binding activity in sucrose gradient sedimentation: alpha with Mr approximately 260,000, beta 1 with Mr approximately 39,000, and beta 2 with Mr approximately 37,000. The alpha subunit, both purified and in intact synaptosomes, is shown to behave anomalously during sodium dodecyl sulfate-polyacrylamide gel electrophoresis exhibiting an unusually high extrapolated electrophoretic free mobility. A subunit stoichiometry of alpha 1(beta 1)1(beta 2)1 is proposed.