Rösgen J, Hinz H J
Institut für Physikalische Chemie, Westfälische Wilhelms-Universität, Schlossplatz 4/7, Münster, 48149, Germany.
J Mol Biol. 2001 Mar 2;306(4):809-24. doi: 10.1006/jmbi.2000.4359.
Heat capacity curves as obtained from differential scanning calorimetry are an outstanding source for molecular information on protein folding and ligand-binding energetics. However, deconvolution of C(p) data of proteins in the presence of ligands can be compromised by indeterminacies concerning the correct choice of the statistical thermodynamic ensemble. By convent, the assumption of constant free ligand concentration has been used to derive formulae for the enthalpy. Unless the ligand occurs at large excess, this assumption is incorrect. Still the relevant ensemble is the grand canonical ensemble. We derive formulae for both constraints, constancy of total or free ligand concentration and illustrate the equations by application to the typical equilibrium Nx <=> N + x <=> D + x. It is demonstrated that as long as the thermodynamic properties of the ligand can be completely corrected for by performing a reference measurement, the grand canonical approach provides the proper and mathematically significantly simpler choice. We demonstrate on the two cases of sequential or independent ligand-binding the fact, that similar binding mechanisms result in different and distinguishable heat capacity equations. Finally, we propose adequate strategies for DSC experiments as well as for obtaining first estimates of the characteristic thermodynamic parameters, which can be used as starting values in a global fit of DSC data.
差示扫描量热法得到的热容曲线是获取蛋白质折叠和配体结合能量学分子信息的重要来源。然而，在存在配体的情况下，蛋白质C(p)数据的反褶积可能会因统计热力学系综正确选择的不确定性而受到影响。按照惯例，一直采用游离配体浓度恒定的假设来推导焓的公式。除非配体大量过量，否则该假设是不正确的。相关的系综仍然是巨正则系综。我们推导了总配体浓度或游离配体浓度恒定这两种约束条件下的公式，并通过应用于典型的平衡Nx <=> N + x <=> D + x来说明这些方程。结果表明，只要通过进行参考测量能够完全校正配体的热力学性质，巨正则方法就是合适且数学上显著更简单的选择。我们在配体顺序结合或独立结合的两种情况下证明，相似的结合机制会导致不同且可区分的热容方程。最后，我们提出了差示扫描量热实验以及获得特征热力学参数初步估计值的适当策略，这些估计值可作为差示扫描量热数据全局拟合的起始值。
