Department of Molecular Organization of Biological Structures, Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow, Russia.
PLoS One. 2013 Sep 10;8(9):e74367. doi: 10.1371/journal.pone.0074367. eCollection 2013.
The methodology for quantification of the anti-aggregation activity of protein and chemical chaperones has been elaborated. The applicability of this methodology was demonstrated using a test-system based on dithiothreitol-induced aggregation of bovine serum albumin at 45°C as an example. Methods for calculating the initial rate of bovine serum albumin aggregation (v agg) have been discussed. The comparison of the dependences of v agg on concentrations of intact and cross-linked α-crystallin allowed us to make a conclusion that a non-linear character of the dependence of v agg on concentration of intact α-crystallin was due to the dynamic mobility of the quaternary structure of α-crystallin and polydispersity of the α-crystallin-target protein complexes. To characterize the anti-aggregation activity of the chemical chaperones (arginine, arginine ethyl ester, arginine amide and proline), the semi-saturation concentration [L]0.5 was used. Among the chemical chaperones studied, arginine ethyl ester and arginine amide reveal the highest anti-aggregation activity ([L]0.5 = 53 and 58 mM, respectively).
已经详细阐述了蛋白质和化学伴侣抗聚集活性的定量方法。该方法的适用性已通过使用基于二硫苏糖醇诱导的牛血清白蛋白在 45°C 下聚集的测试系统为例进行了证明。还讨论了计算牛血清白蛋白聚集初始速率（vagg）的方法。比较 vagg 对完整和交联α-晶体蛋白浓度的依赖性，使我们能够得出结论，vagg 对完整α-晶体蛋白浓度的依赖性的非线性特征是由于α-晶体蛋白的四级结构的动态迁移性和α-晶体蛋白-靶蛋白复合物的多分散性。为了表征化学伴侣（精氨酸、精氨酸乙酯、精氨酸酰胺和脯氨酸）的抗聚集活性，使用半饱和浓度 [L]0.5。在所研究的化学伴侣中，精氨酸乙酯和精氨酸酰胺显示出最高的抗聚集活性（[L]0.5=53 和 58mM，分别）。
