Effect of osmolytes on the exchange rates of backbone amide protons in proteins.
Effect of osmolytes on the exchange rates of backbone amide protons in proteins.
复制标题
渗透剂对蛋白质中骨架酰胺质子交换率的影响。
DOI：
10.1021/bi9712798
复制
发表时间：
1998
期刊：
影响因子：
2.9
通讯作者：
R. Leatherbarrow
中科院分区：
文献类型：
--
作者：
R. Foord;R. Leatherbarrow
渗透剂是所有生物体（除盐细菌外）的细胞在高应激情况（例如极端盐浓度、高温等）下产生的小有机溶质，用于稳定其大分子，从而保持生物活性。它们不直接与大分子相互作用，而是通过改变细胞环境中的溶剂性质来起作用，因此它们的存在间接改变了蛋白质的稳定性。在本文中，我们研究了模型渗透剂甘氨酸对胰凝乳蛋白酶抑制剂 2 和马心细胞色素 c 两种蛋白质稳定性的影响。我们使用核磁共振来监测这种渗透剂对酰胺氢交换率的影响，这使得探针可以在蛋白质结构内的离散点上进行探测。特定主链酰胺质子的氢交换率提供有关局部结构波动的信息，这些波动使这些酰胺暴露于溶剂并允许交换发生。我们发现高浓度甘氨酸渗透剂的存在对所研究的蛋白质具有深远的稳定作用，同时伴随着大多数缓慢交换酰胺质子的交换速率常数的大幅降低。光谱表明，这种情况的发生没有使三维结构发生显着变化。然而，对单一蛋白质内各个酰胺质子的影响并不均匀，并且观察到影响程度的差异很大。其范围从汇率没有明显变化到汇率常数下降超过两个数量级。渗透剂似乎改变了许多不同的过程，这些过程有助于观察到的汇率，并且没有简单的概括可以预测任何单独位置的稳定程度。根据所研究蛋白质的可用结构讨论结果。
Osmolytes are small organic solutes produced by the cells of all organisms (except halobacteria) in high stress situations (e.g. extremes of salt concentration, high temperature, etc.) to stabilize their macromolecules and so conserve biological activity. They do not interact with the macromolecule directly but act by altering the solvent properties in the cellular environment, and so their presence indirectly modifies the stability of proteins. In this paper we examine the effect of a model osmolyte, glycine, on the stabilization of two proteins, chymotrypsin inhibitor 2 and horse heart cytochrome c. We have used NMR to monitor the effect of this osmolyte on amide hydrogen exchange rates, which allows a probe at discrete points within the protein structure. Hydrogen exchange rates of specific backbone amide protons provide information about the localized structural fluctuations that expose these amides to solvent and allow exchange to take place. We find that the presence of a high concentration of glycine osmolyte has a profound stabilizing effect on the proteins studied, which is accompanied by a large reduction of the exchange rate constants of most slowly exchanging amide protons. The spectra indicate that this arises without significant changes in the three-dimensional structure. However, the effects on individual amide protons within a single protein were not uniform, and a wide variation in the magnitude of the effects was observed. This ranged from no apparent change in the exchange rate, to decreases in the exchange rate constant by over 2 orders of magnitude. The osmolyte appears to alter a number of different processes that contribute to the observed exchange rates, and no simple generalization allows prediction of the extent of stabilization at any individual location. The results are discussed in light of the available structures of the proteins studied.