Correlations between the charge of proteins and the number of ionizable groups they incorporate: Studies using protein charge ladders, capillary electrophoresis, and Debye-Huckel theory
Correlations between the charge of proteins and the number of ionizable groups they incorporate: Studies using protein charge ladders, capillary electrophoresis, and Debye-Huckel theory
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DOI：
10.1021/ja991526q
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发表时间：
1999-11-24
影响因子：
15
通讯作者：
Whitesides, GM
中科院分区：
文献类型：
--
作者：
Carbeck, JD;Colton, IJ;Whitesides, GM
用毛细管电泳法测定了牛碳酸酐酶Ⅱ、人碳酸酐酶Ⅱ、细胞色素c、溶菌酶、超氧化物歧化酶、卵清蛋白及其衍生物部分中和赖氨酸氨基和/或天冬氨酸和谷氨酸羧基所产生的电泳率。对于净电荷总量最小(正负)的这些蛋白质的衍生物，它们的电荷值和CE、Z(CE)测得的电荷值与中性导数的带电基数n呈线性相关。对于具有较大净电荷值的这些蛋白质的衍生物，它们的mU(电性)和Z(CE)值与n呈非线性相关。本工作中的几个观察结果表明，这些蛋白质上可电离基团的pK(A)值的变化可能是造成这种非线性相关的原因。用德拜-休克尔理论计算了所有6种蛋白质的导数表面的静电势，并计算了它们的电势。将这些值与通过将电荷分配给每个蛋白质与n成正比而计算出的静电电位值绘制成曲线图。所有六种蛋白质的数据都沿着一条共同的曲线下降，而不考虑电泳液中一价阳离子的浓度。
The values of electrophoretic mobility, mu(electro), of bovine carbonic anhydrase II, human carbonic anhydrase II, cytochrome c, lysozyme, superoxide dismutase, ovalbumin, and derivatives of these proteins produced by partial neutralization of Lys epsilon-NH3+ and/or Asp and Glu carboxyl groups were measured using capillary electrophoresis (CE). For derivatives of these proteins with the lowest overall values of net charge (either positive or negative), the values of mu(electro) and the values of charge measured by CE, Z(CE), demonstrate a linear correlation with the number of charged groups, n, converted to neutral derivatives. For derivatives of these proteins with larger values of net charge, the values of mu(electro) and Z(CE) demonstrate a nonlinear correlation with n. Several observations made in this work suggest that shifts in the values of pk(a) of the ionizable groups on these proteins likely contribute to the observed nonlinear correlation. Debye-Huckel theory was used to calculate values of electrostatic potential at the surface of the derivatives of all six proteins from the measured values of mu(electro). These values were plotted against the values of electrostatic potential calculated by assigning a charge to each protein in direct proportion to n. The data for all six proteins fell along a single common curve, regardless of the concentration of monovalent cations in the electrophoresis buffer.