Characterization of globular protein solutions by dynamic light scattering, electrophoretic mobility, and viscosity measurements
Characterization of globular protein solutions by dynamic light scattering, electrophoretic mobility, and viscosity measurements
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DOI：
10.1021/la800548p
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发表时间：
2008-07-01
期刊：
影响因子：
3.9
通讯作者：
Adamczyk, Zbigniew
中科院分区：
文献类型：
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作者：
Jachimska, Barbara;Wasilewska, Monika;Adamczyk, Zbigniew
在这项工作中，两个球状蛋白质-牛血清白蛋白（BSA）具有67 kDa的分子量和人血清白蛋白（HSA）具有69 kDa的分子量的物理化学性质的特点。这些蛋白质的体积特性涉及的扩散系数（流体动力学半径），电泳迁移率，和动态粘度作为蛋白质溶液浓度的函数为各种pH值。流体动力学半径的数据表明，协会的蛋白质分子，最有可能形成紧凑的二聚体。使用流体动力学直径和电泳迁移率数据允许确定蛋白质表面上的未补偿（电动）电荷的数量。将电泳迁移率数据转换为zeta电位值，从而可以确定这些蛋白质的等电点（iep）。根据先前的实验数据和来自氨基酸组成和pK值的理论估计，发现这两种蛋白质的pH为5.1。为了进一步确定蛋白质溶液的稳定性，进行动态粘度测量，作为它们在各种pH值下的体积浓度的函数。从这些测量得到的特性粘度解释的布伦纳模型，这是适用于硬球状颗粒。发现当假设蛋白质尺寸为9.5 x 5 x 5 nm（3）（长球体）时，这些蛋白质的特性粘度的实验值与该模型非常一致。通过这些测量排除了形成缔合度高于2的线性聚集体的可能性。得出的结论是，动态粘度和动态光散射的组合可以被利用作为一个方便的工具，用于检测不仅在悬浮液中的蛋白质聚集的发病，而且这些聚集体的形式和组成。
In this work, physicochemical properties of two globular proteins-bovine serum albumin (BSA) having a molecular weight of 67 kDa and human serum albumin (HSA) having a molecular weight of 69 kDa-were characterized. The bulk characteristics of these proteins involved the diffusion coefficient (hydrodynamic radius), electrophoretic mobility, and dynamic viscosity as a function of protein solution concentration for various pH values. The hydrodynamic radius data suggested an association of protein molecules, most probably forming compact dimers. Using the hydrodynamic diameter and the electropheretic mobility data allowed the determination of the number of uncompensated (electrokinetic) charges on protein surfaces. The electrophoretic mobility data were converted to zeta potential values, which allowed one to determine the isoelectric point (iep) of these proteins. It was found to be at pH 5.1 for both proteins, in accordance with previous experimental data and theoretical estimations derived from amino acid composition and pK values. To determine further the stability of protein solutions, dynamic viscosity measurements were carried out as a function of their bulk volume concentration for various pH values. The intrinsic viscosity derived from these measurements was interpreted in terms of the Brenner model, which is applicable to hard spheroidal particles. It was found that the experimental values of the intrinsic viscosity of these proteins were in good agreement with this model when assuming protein dimensions of 9.5 x 5 x 5 nm(3) (prolate spheroid). The possibility of forming linear aggregates of association degree higher than 2 was excluded by these measurements. It was concluded that the combination of dynamic viscosity and dynamic light scattering can be exploited as a convenient tool for detecting not only the onset of protein aggregation in suspensions but also the form and composition of these aggregates.