Structural and folding dynamic properties of the T70N variant of human lysozyme*
Structural and folding dynamic properties of the T70N variant of human lysozyme*
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DOI：
10.1074/jbc.m211000200
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发表时间：
2003-07-11
影响因子：
4.8
通讯作者：
Bellotti, V
中科院分区：
文献类型：
--
作者：
Esposito, G;Garcia, J;Bellotti, V
通过对人溶菌酶的两种淀粉样蛋白变体I56 T和D 67 H进行的生物化学和生物物理学研究，大大提高了从天然球状蛋白到纤维状聚合物的过渡机制的定义。在这里，我们报告的热力学和动力学数据的折叠以及结构特征的自然发生的变种人溶菌酶，T70 N，这是目前在英国人口中的等位基因频率为5%，根据临床和组织病理学数据，是不是淀粉样蛋白。这种变体比野生型蛋白稳定性低3.7 kcal/mol，但比病理性淀粉样蛋白变体更稳定。胍中的解折叠动力学比野生型快6倍，但比淀粉样变形体慢3倍和20倍。与野生型相比，酶催化参数，如最大速度和亲和力降低。的解决方案的结构，确定由H-1 NMR和建模计算，表现出更紧凑的安排之间的界面上的β-折叠结构域和随后的环的一侧和部分的α结构域的另一侧，与野生型蛋白质。这与淀粉样蛋白生成变体D 67 H所示的构象变化相反，但它解释了T70 N的稳定性和催化性能降低。
Definition of the transition mechanism from the native globular protein into fibrillar polymer was greatly improved by the biochemical and biophysical studies carried out on the two amyloidogenic variants of human lysozyme, I56T and D67H. Here we report thermodynamic and kinetic data on folding as well as structural features of a naturally occurring variant of human lysozyme, T70N, which is present in the British population at an allele frequency of 5% and, according to clinical and histopathological data, is not amyloidogenic. This variant is less stable than the wild-type protein by 3.7 kcal/mol, but more stable than the pathological, amyloidogenic variants. Unfolding kinetics in guanidine are six times faster than in the wild-type, but three and twenty times slower than in the amyloidogenic variants. Enzyme catalytic parameters, such as maximal velocity and affinity, are reduced in comparison to the wild-type. The solution structure, determined by H-1 NMR and modeling calculations, exhibits a more compact arrangement at the interface between the beta-sheet domain and the subsequent loop on one side and part of the alpha domain on the other side, compared with the wild-type protein. This is the opposite of the conformational variation shown by the amyloidogenic variant D67H, but it accounts for the reduced stability and catalytic performance of T70N.