Atomic resolution structures of ribonuclease A at six pH values
Atomic resolution structures of ribonuclease A at six pH values
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DOI：
10.1107/s0907444901021758
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发表时间：
2002-03-01
期刊：
影响因子：
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通讯作者：
Mazzarella, L
中科院分区：
文献类型：
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作者：
Berisio, R;Sica, F;Mazzarella, L
蛋白质晶体的衍射图延伸到原子分辨率，确保了分子结构的非常准确的图像，并使研究与蛋白质功能相关的微妙现象成为可能。在pH分别为5.2、5.9、6.3、7.1、8.0和8.8，分辨极限为1.05-1.15埃的条件下，对牛胰腺核糖核酸酶的六种结构进行了精制。这里描述了六种结构和几个方面的总体描述，主要是关于pH触发的构象变化。由于预计会有细微的变化，因此首先对六个改进后的模型进行了彻底的验证评估。一些立体化学参数，如N-C-α-C角和羰基C原子的锥化，表明通常用于精炼的标准目标值及其权重可能需要修订。对这六种结构的详细比较为Lys41在催化中的作用提供了实验证据。此外，对与活性中心中模拟RNA磷酸基团的硫酸盐阴离子的pH依赖结合有关的结构效应也给出了见解。最后，这些结果支持了一些关于Cys65和Cys72之间的二硫键在RNaseA折叠中的作用的热力学和动力学实验数据。
The diffraction pattern of protein crystals extending to atomic resolution guarantees a very accurate picture of the molecular structure and enables the study of subtle phenomena related to protein functionality. Six structures of bovine pancreatic ribonuclease at the pH* values 5.2, 5.9, 6.3, 7.1, 8.0 and 8.8 and at resolution limits in the range 1.05-1.15 Angstrom have been refined. An overall description of the six structures and several aspects, mainly regarding pH-triggered conformational changes, are described here. Since subtle variations were expected, a thorough validation assessment of the six refined models was first carried out. Some stereochemical parameters, such as the N-C-alpha-C angle and the pyramidalization at the carbonyl C atoms, indicate that the standard target values and their weights typically used in refinement may need revision. A detailed comparison of the six structures has provided experimental evidence on the role of Lys41 in catalysis. Furthermore, insights are given into the structural effects related to the pH-dependent binding of a sulfate anion, which mimics the phosphate group of RNA, in the active site. Finally, the results support a number of thermodynamic and kinetic experimental data concerning the role of the disulfide bridge between Cys65 and Cys72 in the folding of RNase A.