Ligand-induced biphasic protein denaturation.
Ligand-induced biphasic protein denaturation.
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DOI：
10.1016/s0021-9258(19)34083-9
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发表时间：
1990-03
期刊：
影响因子：
--
通讯作者：
A. Shrake;P. Ross
中科院分区：
文献类型：
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作者：
A. Shrake;P. Ross
的热力学计算的结果是基于实验观察，并结合了配体结合的两个状态，热变性的蛋白质的影响的过剩的热容量。对于一个单一的结合位点上的天然物种和在亚饱和浓度的配体，双峰或单峰热谱图计算仅仅通过假设一个较大或较小的配体缔合常数，分别。该简化情况的计算热谱图显示了在不存在和存在蛋白质对其具有较高、中等和较低亲和力的三种配体的情况下通过差示扫描量热法对脱脂人白蛋白单体观察到的那些显著特征（Shrake，A.，还有罗斯，警察。（1988）J.Biol.Chem.263，15392-15399）。计算表明，双相解折叠可以导致在解折叠过程中由于释放的结合配体的蛋白质变性引起的游离配体浓度的大幅增加，变性的自由能（和转变温度）的显着增加。这种配体诱导的双相变性不涉及大分子的亚结构，但来自扰动，在展开过程中，配体结合平衡，这是耦合到折叠和未折叠的蛋白质物种之间的平衡。因此，这种双峰性并不限于热诱导的解折叠，而是独立于用于影响变性的手段而起作用，因此在研究配体存在下的任何大分子折叠/解折叠反应时必须考虑。
The results of a thermodynamic calculation of the excess heat capacity that is based on experimental observations and that incorporates the effects of ligand binding on the two-state, thermal denaturation of a protein are presented. For a protein with a single-binding site on the native species and at subsaturating concentrations of ligand, bimodal or unimodal thermograms were computed merely by assuming a larger or smaller ligand association constant, respectively. The calculated thermograms for this simplified case show the salient features of those observed by differential scanning calorimetry for defatted human albumin monomer in the absence and presence of three ligands for which the protein has higher, intermediate, and lower affinity (Shrake, A., and Ross, P. D. (1988) J. Biol. Chem. 263, 15392-15399). The computation demonstrates that biphasic unfolding can result from a significant increase in the free energy of denaturation (and the transition temperature) during the course of unfolding due to a substantial increase in free ligand concentration caused by the release of bound ligand by denaturing protein. Such ligand-induced biphasic denaturation does not relate to macromolecular substructure but derives from a perturbation, during unfolding, of the ligand binding equilibrium, which is coupled to the equilibrium between the folded and unfolded protein species. Thus, this bimodality is not limited to thermally induced unfolding but is operative independent of the means used to effect denaturation and therefore must be considered when studying any macromolecular folding/unfolding reaction in the presence of ligand.