Improvement of the solubilization of proteins in two-dimensional electrophoresis with immobilized pH gradients
Improvement of the solubilization of proteins in two-dimensional electrophoresis with immobilized pH gradients
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DOI：
10.1002/elps.1150180303
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发表时间：
1997-03-01
期刊：
影响因子：
2.9
通讯作者：
Lunardi, J
中科院分区：
文献类型：
--
作者：
Rabilloud, T;Adessi, C;Lunardi, J
用高分辨率双向凝胶电泳法分离了溶解性差的膜蛋白和核蛋白。等电聚焦和固定的pH梯度导致蛋白质在所得到的二维图谱中的严重定量损失，尽管分辨率通常很高。使用含有各种洗涤剂和杂色剂的变性溶液可以改善蛋白质的溶解性。用含有尿素、硫脲和洗涤剂(非离子和两性离子)的变性溶液效果最好。含有硫脲的变性混合物对微粒体和核蛋白以及微管蛋白都很有用，微管蛋白是一种高度容易聚集的蛋白质。
Membrane and nuclear proteins of poor solubility have been separated by high resolution two-dimensional (2-D) gel electrophoresis. Isoelectric focusing with immobilized pH gradients leads to severe quantitative losses of proteins in the resulting 2-D map, although the resolution is usually high. Protein solubility could be improved by using denaturing solutions containing various detergents and chaotropes. Best results were obtained with a denaturing solution containing urea, thiourea,and detergents (both nonionic and zwitterionic). The usefulness of thiourea-containing denaturing mixtures is shown for microsomal and nuclear proteins as well as for tubulin, a protein highly prone to aggregation.