DETERMINATION OF THE EFFECTIVE CHARGE OF A PROTEIN IN SOLUTION BY CAPILLARY ELECTROPHORESIS
DETERMINATION OF THE EFFECTIVE CHARGE OF A PROTEIN IN SOLUTION BY CAPILLARY ELECTROPHORESIS
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DOI：
10.1073/pnas.91.25.12027
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发表时间：
1994-12-06
影响因子：
11.1
通讯作者：
WHITESIDES, GM
中科院分区：
文献类型：
--
作者：
GAO, JM;GOMEZ, FA;WHITESIDES, GM
本文介绍了用毛细管电泳法测定溶液中蛋白质有效电荷的两种方法，并以具有代表性的蛋白质为例进行了验证。一种方法是用4-巯基异硫氰酸酯或乙酸酐对赖氨酸的ε -氨基进行共价修饰，从而产生“电荷阶梯”——一种蛋白质的一系列衍生物，其电荷增量已知不同，但在水动力阻力方面差异极小。在第二种方法中，通过蛋白质与不同带电配体的非共价结合产生等效的电荷阶梯。分析蛋白质及其衍生物的电泳迁移率作为附加电荷的函数，可以估计未修饰蛋白质的有效电荷。这种类型的分析允许在不知道其组成、结构或氨基酸序列的情况下估计蛋白质的有效电荷。
This paper describes two methods to estimate the effective charge of a protein in solution by capillary electrophoresis and demonstrates these methods by using representative proteins. In one method, a ''charge ladder''-a series of derivatives of a protein differing by known increments of charge but differing only minimally in hydrodynamic drag-is generated by covalent modification of the epsilon-amino groups of lysines with 4-sulfophenyl isothiocyanate or acetic anhydride. In the second method, the equivalent of a charge ladder is produced by noncovalent association of a protein with differently charged ligands. Analysis of the electrophoretic mobilities of the protein and its derivatives as a function of added charge allows the effective charge to be estimated for the unmodified protein. This type of analysis permits estimation of the effective charge of a protein without knowing its composition, structure, or amino acid sequence.