Jagiellonian University, Faculty of Biochemistry, Biophysics and Biotechnology, Department of Physical Biochemistry, Gronostajowa 7, 30-387 Kraków, Poland.
Int J Biol Macromol. 2018 Dec;120(Pt A):128-134. doi: 10.1016/j.ijbiomac.2018.08.012. Epub 2018 Aug 8.
In proteins, proton exchange is caused due to the changes in the proton affinity (pK) of ionizable groups that are engaged in conformational changes induced by the binding of a ligand. In addition, knowledge regarding the type and number of such ionizable groups is very important to understand the pH-dependent changes of the thermodynamic parameters. Therefore, in this study, we performed a comprehensive analysis of proton exchange by using beta lactoglobulin (Blg), a representative of the lipocalin family of proteins. We used isothermal titration calorimetry to evaluate the proton exchange during binding with sodium dodecyl sulfate (SDS) at different pHs ranging from 2 to 9. SDS binds to Blg in all studied range of pHs and enthalpy-driven reactions are observed in acidic pH, whereas enthalpy-entropy driven reactions are observed in neutral and basic pHs. Enthalpy-entropy compensation leads to relatively small changes in the association constants with the maximal value at pH = 8.0. The simultaneous analysis of the number of exchanged protons, the binding constants, and the enthalpy was performed in the range pH 5.5-9. At least 4 residues that change their ionization pattern are needed to explain the observed pH dependence.
在蛋白质中，质子交换是由于参与构象变化的可离子化基团的质子亲和度（pK）发生变化而引起的，这些构象变化是由配体结合引起的。此外，了解这类可离子化基团的类型和数量对于理解热力学参数随 pH 的变化非常重要。因此，在这项研究中，我们使用β乳球蛋白（Blg）作为代表性的亲脂素家族蛋白质，进行了全面的质子交换分析。我们使用等温滴定量热法在 pH 值为 2 至 9 的不同范围内评估与十二烷基硫酸钠（SDS）结合过程中的质子交换。SDS 在所有研究的 pH 值范围内与 Blg 结合，在酸性 pH 值下观察到焓驱动反应，而在中性和碱性 pH 值下观察到焓熵驱动反应。焓熵补偿导致结合常数相对较小的变化，最大值出现在 pH=8.0。在 pH 5.5-9 的范围内，同时分析了交换质子的数量、结合常数和焓。至少需要 4 个残基来改变它们的离子化模式，才能解释观察到的 pH 依赖性。
