通过毛细管电泳法测定溶液中蛋白质的有效电荷
Determination of the effective charge of a protein in solution by capillary electrophoresis.
作者信息
Gao J, Gomez F A, Härter R, Whitesides G M
机构信息
Department of Chemistry, Harvard University, Cambridge, MA 02138.
出版信息
Proc Natl Acad Sci U S A. 1994 Dec 6;91(25):12027-30. doi: 10.1073/pnas.91.25.12027.
Abstract
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.
摘要
本文介绍了两种通过毛细管电泳估算溶液中蛋白质有效电荷的方法，并使用代表性蛋白质对这些方法进行了验证。在一种方法中，通过用4-磺基苯基异硫氰酸酯或乙酸酐对赖氨酸的ε-氨基进行共价修饰，生成“电荷梯”——一系列电荷增量已知但流体动力学阻力差异极小的蛋白质衍生物。在第二种方法中，通过蛋白质与带不同电荷的配体非共价结合产生等效的电荷梯。分析蛋白质及其衍生物的电泳迁移率随添加电荷的变化情况，可以估算未修饰蛋白质的有效电荷。这种分析类型允许在不知道蛋白质组成、结构或氨基酸序列的情况下估算其有效电荷。
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