Protein-protein interactions in dilute to concentrated solutions: α-chymotrypsinogen in acidic conditions.
Protein-protein interactions in dilute to concentrated solutions: α-chymotrypsinogen in acidic conditions.
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DOI：
10.1021/jp412301h
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发表时间：
2014-06-05
期刊：
影响因子：
--
通讯作者：
Roberts CJ
中科院分区：
文献类型：
--
作者：
Blanco MA;Perevozchikova T;Martorana V;Manno M;Roberts CJ
通过静态光散射（SLS）和动态光散射（DLS）以及小角中子散射（SANS）研究了α-凝乳胰蛋白酶原蛋白-蛋白相互作用与酸性条件下蛋白质和盐浓度的关系。通过Kirkwood-Buff积分G22和SLS和SANS数据中的静态结构因子S(q)来探测净蛋白-蛋白相互作用。G22是通过使用局部泰勒级数方法将瑞利比与蛋白质浓度进行回归得到的，该方法不需要假设分子间相互作用的潜在形式或性质。此外，采用拟合有效相互作用势的传统方法进一步分析G22和S(q)。虽然拟合的模型参数在物理上并不总是真实的，但G22和S的数值（q→0）作为蛋白质浓度的函数与SLS和SANS非常一致。在稀释状态下，G22的拟合值与渗透二阶维里系数B22的拟合值一致，表明静电相互作用是α-凝乳胰蛋白酶原溶液中胶体相互作用的主要贡献。然而，随着蛋白质浓度的增加，蛋白质-蛋白质相互作用的强度降低，在低盐浓度下下降更为明显。结果与有效的“拥挤”或排除体积对G22的贡献是一致的，因为即使在这里使用的蛋白质浓度的中等范围内（<40 g/L），远程静电斥力也很突出。通过将DLS数据的集体扩散系数测量值与S（q→0）测量值相结合，评估水动力因子H(q→0)，证实了这些明显的拥挤效应，并对其进行了量化。H（q→0）明显小于相应的硬球体系，这表明如果仅使用DLS来评估蛋白质相互作用，水动力非理想性可能导致关于B22、G22和静态蛋白质-蛋白质相互作用的定性错误结论。
Protein–protein interactions were investigated for α-chymotrypsinogen by static and dynamic light scattering (SLS and DLS, respectively), as well as small-angle neutron scattering (SANS), as a function of protein and salt concentration at acidic conditions. Net protein–protein interactions were probed via the Kirkwood–Buff integral G22 and the static structure factor S(q) from SLS and SANS data. G22 was obtained by regressing the Rayleigh ratio versus protein concentration with a local Taylor series approach, which does not require one to assume the underlying form or nature of intermolecular interactions. In addition, G22 and S(q) were further analyzed by traditional methods involving fits to effective interaction potentials. Although the fitted model parameters were not always physically realistic, the numerical values for G22 and S(q → 0) were in good agreement from SLS and SANS as a function of protein concentration. In the dilute regime, fitted G22 values agreed with those obtained via the osmotic second virial coefficient B22 and showed that electrostatic interactions are the dominant contribution for colloidal interactions in α-chymotrypsinogen solutions. However, as protein concentration increases, the strength of protein–protein interactions decreases, with a more pronounced decrease at low salt concentrations. The results are consistent with an effective “crowding” or excluded volume contribution to G22 due to the long-ranged electrostatic repulsions that are prominent even at the moderate range of protein concentrations used here (<40 g/L). These apparent crowding effects were confirmed and quantified by assessing the hydrodynamic factor H(q → 0), which is obtained by combining measurements of the collective diffusion coefficient from DLS data with measurements of S(q → 0). H(q → 0) was significantly less than that for a corresponding hard-sphere system and showed that hydrodynamic nonidealities can lead to qualitatively incorrect conclusions regarding B22, G22, and static protein–protein interactions if one uses only DLS to assess protein interactions.
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