Carra J H, Anderson E A, Privalov P L
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
Protein Sci. 1994 Jun;3(6):952-9. doi: 10.1002/pro.5560030610.
Staphylococcal nuclease, at low pH and in the presence of high salt concentrations, has previously been proposed to exist in a partially folded or molten globule form called the "A-state" (Fink et al., 1993, Protein Sci 2:1155-1160). We have found that the A-state of nuclease at pH 2.1 in the presence of moderate to high salt concentrations and at low temperature exists in a substantially folded form structurally more similar to a native state. The A-state has the far-UV circular dichroism spectra characteristic of the native protein, which indicates that it has a large degree of secondary structure. Upon heating, the A-state denatures with a sigmoidal change in far-UV ellipticity and an observable peak in a differential scanning calorimeter trace, indicating that it is thermodynamically distinct from the denatured state. Three different mutations in a residue normally buried in the protein's core stabilize or destabilize the A-state in the same way as they affect the denaturation of the native state. The A-state must, therefore, contain at least some tertiary packing of side chains. Unlike the native state, which shows cold denaturation at low temperatures, the A-state is most stable at temperatures below 0 degrees C.
以前有人提出，在低pH值和高盐浓度存在的情况下，葡萄球菌核酸酶以一种部分折叠或熔融球状的形式存在，称为“A态”（芬克等人，1993年，《蛋白质科学》2：1155 - 1160）。我们发现，在中等至高盐浓度存在且低温条件下，pH 2.1时核酸酶的A态以一种结构上与天然态更相似的基本折叠形式存在。A态具有天然蛋白质特有的远紫外圆二色光谱，这表明它具有很大程度的二级结构。加热时，A态变性，远紫外椭圆率呈S形变化，差示扫描量热仪曲线上出现一个可观察到的峰，表明它在热力学上与变性态不同。通常埋藏在蛋白质核心中的一个残基发生的三种不同突变，对A态的稳定或不稳定作用，与它们对天然态变性的影响方式相同。因此，A态必须至少包含一些侧链的三级堆积。与在低温下显示冷变性的天然态不同，A态在低于0摄氏度的温度下最稳定。
