The Drosophila acetylcholine receptor subunit D alpha5 is part of an alpha-bungarotoxin binding acetylcholine receptor.
The Drosophila acetylcholine receptor subunit D alpha5 is part of an alpha-bungarotoxin binding acetylcholine receptor.
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果蝇乙酰胆碱受体亚基 D α5 是 α-银环蛇毒素结合乙酰胆碱受体的一部分。
DOI：
10.1074/jbc.m409639200
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Schmidt-Glenewinkel,Thomas
中科院分区：
文献类型：
--
作者：
Wu,Peipei;Ma,Dongdong;Pierzchala,Marek;Wu,Jun;Yang,Lee-Chuan;Mai,Xiaoping;Chang,Xiaoying;Schmidt-Glenewinkel,Thomas
黑腹果蝇的中枢神经系统含有α-班加罗毒素结合蛋白，具有烟碱乙酰胆碱受体的特性。该蛋白从果蝇前体制备的膜中纯化了5800倍。蛋白用1% Triton X-100和0.5 m氯化钠溶解，α-蛇毒素柱和扁豆凝集素亲和柱纯化。纯化蛋白的比活性为3.9 μmol /g蛋白的125i -α-虫毒结合位点。用十二烷基硫酸钠聚丙烯酰胺凝胶电泳法测定了纯化后受体的亚基组成。该亚基谱与利用125i -α-兔毒的可光解甲基-4-叠氮苯并咪酯衍生物对膜结合蛋白进行情景标记所显示的亚基谱相同。纯化后的受体显示两个不同的蛋白带，分子量分别为42和57 kDa。通过对纯化蛋白复合物在H2O和D2O中的沉降分析和凝胶过滤，计算出其质量为270 kDa。该受体的直径为20米，直径为9.4米，斯托克半径为7.4纳米。计算摩擦系数为1.7，表明高度不对称的蛋白复合物与跨膜蛋白兼容，形成离子通道。经胰蛋白酶消化42 kda蛋白后获得的肽序列可以通过序列比较特异性鉴定果蝇ad α5亚基。针对Dα5亚基的肽特异性抗体进一步证明该亚基是来自果蝇中枢神经系统的α-虫毒结合烟碱乙酰胆碱受体的组成部分。
The central nervous system ofDrosophila melanogastercontains an α-bungarotoxin-binding protein with the properties expected of a nicotinic acetylcholine receptor. This protein was purified 5800-fold from membranes prepared fromDrosophilaheads. The protein was solubilized with 1% Triton X-100 and 0.5msodium chloride and then purified using an α-cobratoxin column followed by a lentil lectin affinity column. The purified protein had a specific activity of 3.9 μmol of125I-α-bungarotoxin binding sites/g of protein. The subunit composition of the purified receptor was determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. This subunit profile was identical with that revealed byin situlabeling of the membrane-bound protein using the photolyzable methyl-4-azidobenzoimidate derivative of125I-α-bungarotoxin. The purified receptor reveals two different protein bands with molecular masses of 42 and 57 kDa. From sedimentation analysis of the purified protein complex in H2O and D2O and gel filtration, a mass of 270 kDa was calculated. The receptor has as20,wof 9.4 and a Stoke's radius of 7.4 nm. The frictional coefficient was calculated to be 1.7 indicating a highly asymmetric protein complex compatible with a transmembrane protein forming an ion channel. The sequence of a peptide obtained after tryptic digestion of the 42-kDa protein allowed the specific identification of theDrosophilaDα5 subunit by sequence comparison. A peptide-specific antibody raised against the Dα5 subunit provides further evidence that this subunit is a component of an α-bungarotoxin binding nicotinic acetylcholine receptor from the central nervous system ofDrosophila.