pH effects on the stability and dimerization of procaspase-3.
pH effects on the stability and dimerization of procaspase-3.
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pH 对 procaspase-3 稳定性和二聚化的影响。
DOI：
10.1110/ps.041003305
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Clark,AClay
中科院分区：
文献类型：
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作者：
Bose,Kakoli;Clark,AClay
已知pH依赖性构象变化发生在二聚体procaspase-3中，并且它们已被证明会影响自动饱和的速率。我们研究了半胱氨酸天冬氨酸蛋白酶原-3（C163 S）的平衡解折叠随pH值（pH 8.5至pH 4之间）的变化，以便在折叠和稳定性的背景下检查这些变化。数据显示，在低于pH 5时，半胱氨酸天冬氨酸蛋白酶原二聚体经历pH依赖性解离，使得蛋白质在pH 4时大部分是单体。与此一致，二聚体在pH 8.5和pH 4.75之间通过四态过程展开，其中天然二聚体异构化为二聚体中间体，二聚体中间体解离为单体，然后展开。与此相反，一个小的蛋白质浓度依赖性，观察圆二色性，但不是由荧光发射，在pH 4.5和pH 4.2。在pH 4下收集的数据不存在蛋白浓度依赖性。总体而言，结果是一致的重新分配的人口的天然二聚体（N2）二聚体中间体（I2）的单体中间体（I），作为pH值降低，使在pH值4，“天然”合奏类似于在较高的pH值展开过程中观察到的单体中间体（I）。一个新兴的图片的单体半胱氨酸天冬氨酸蛋白酶原进行了讨论。半胱天冬酶原-3在pH = 7（24-26 kcal/mol）时最稳定，虽然稳定性随pH值降低，但观察到二聚化贡献了大部分（>70%）构象自由能。
pH‐dependent conformational changes are known to occur in dimeric procaspase‐3, and they have been shown to affect the rate of automaturation. We studied the equilibrium unfolding of procaspase‐3(C163S) as a function of pH (between pH 8.5 and pH 4) in order to examine these changes in the context of folding and stability. The data show that the procaspase dimer undergoes a pH‐dependent dissociation below pH 5, so that the protein is mostly monomeric at pH 4. Consistent with this, the dimer unfolds via a four‐state process between pH 8.5 and pH 4.75, in which the native dimer isomerizes to a dimeric intermediate, and the dimeric intermediate dissociates to a monomer, which then unfolds. In contrast, a small protein concentration dependence was observed by circular dichroism, but not by fluorescence emission, at pH 4.5 and pH 4.2. There was no protein‐concentration dependence to the data collected at pH 4. Overall, the results are consistent with the redistribution of the population of native dimer (N2) to dimeric intermediate (I2) to monomeric intermediate (I), as the pH is lowered so that at pH 4, the “native” ensemble resembles the monomeric intermediate (I) observed during unfolding at higher pH. An emerging picture of the monomeric procaspase is discussed. Procaspase‐3 is most stable at pH ∼7 (24–26 kcal/mol), and while the stability decreased with pH, it was observed that dimerization contributes the majority (>70%) of the conformational free energy.