Sequential sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reversed-phase chromatography of unfolded proteins.
Sequential sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reversed-phase chromatography of unfolded proteins.
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未折叠蛋白的连续十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和反相色谱。
DOI：
10.1016/0003-2697(89)90657-x
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发表时间：
1989
影响因子：
2.9
通讯作者：
J. Vanfleteren
中科院分区：
文献类型：
--
作者：
J. Vanfleteren
连续的十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶电泳和反相高效液相色谱在氟碳填料（聚F柱，杜邦公司）提供完全变性，但高度纯化的蛋白质，这是免费的低分子量的物质，并直接进行结构分析。对四种测试蛋白（牛血清白蛋白、碳酸酐酶、肌红蛋白、细胞色素c）印迹到Immobilon膜上的制备洗脱条件进行了优化。用50 mm Tris-HCl（pH 8.25）饱和并含有2% SDS和1% 2-巯基乙醇的苯酚能够吸附已印迹到Immobilon膜上、用考马斯R-250染色并以干片形式储存的蛋白质，而与其分子量基本无关。暴露于低pH值下不降解的多肽也可以用70%甲酸直接从染色的凝胶中有效提取。如果不需要进一步分离多肽，则将溶于1-2% SDS的蛋白质样品在polyF柱上简单运行将有效去除低分子量物质，包括SDS。
Sequential sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and reversed-phase high performance liquid chromatography on a fluorocarbon packing (poly F column, DuPont) provide fully denatured but highly purified protein, which is free of low-molecular-weight substances and directly amenable to structural analysis. Conditions for the preparative elution of four test proteins (bovine serum albumin, carbonic anhydrase, myoglobin, cytochrome c) blotted to Immobilon membranes have been optimized. Phenol saturated with 50 mm Tris-HCl (pH 8.25) and containing 2% SDS and 1% 2-mercaptoethanol is able to elute proteins that have been blotted to Immobilon membranes, stained with Coomassie R-250, and stored as dry sheets, largely irrespective of their molecular mass. Polypeptides that are not degraded by exposure to low pH can also be efficiently extracted directly from stained gels with 70% formic acid. If further separation of polypeptides is not needed, a simple run of a protein sample dissolved in 1–2% SDS on the poly F column will efficiently remove low-molecular-weight substances, including SDS.
通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分离后，从硝酸纤维素膜上制备性洗脱蛋白质。
影响因子：
2.9
作者：
Parekh,BS;Mehta,HB;West,MD;Montelaro,RC
通讯作者：
Montelaro,RC
影响因子：
2.9
作者：
HAGER, DA;BURGESS, RR
通讯作者：
BURGESS, RR