Tehei Moeava, Zaccai Giuseppe
Institut Laue-Langevin, Grenoble, France.
FEBS J. 2007 Aug;274(16):4034-43. doi: 10.1111/j.1742-4658.2007.05953.x.
Work on the relationship between hyperthermophile protein dynamics, stability and activity is reviewed. Neutron spectroscopy has been applied to measure and compare the macromolecular dynamics of various hyperthermophilic and mesophilic proteins, under different conditions. First, molecular dynamics have been analyzed for the hyperthermophile malate dehydrogenase from Methanococcus jannaschii and a mesophilic homologue, the lactate dehydrogenase from Oryctolagus cunniculus (rabbit) muscle. The neutron scattering approach has provided independent measurements of the global flexibility and structural resilience of each protein, and it has been demonstrated that macromolecular dynamics represents one of the molecular mechanisms of thermoadaptation. The resilience was found to be higher for the hyperthermophilic protein, thus ensuring similar flexibilities in both enzymes at their optimal activity temperature. Second, the neutron method has been developed to quantify the average macromolecular flexibility and resilience within the natural crowded environment of the cell, and mean macromolecular motions have been measured in vivo in psychrophile, mesophile, thermophile and hyperthermophile bacteria. The macromolecular resilience in bacteria was found to increase with adaptation to high temperatures, whereas flexibility was maintained within narrow limits, independent of physiological temperature for all cells in their active state. Third, macromolecular motions have been measured in free and immobilized dihydrofolate reductase from Escherichia coli. The immobilized mesophilic enzyme has increased stability and decreased activity, so that its properties are changed to resemble those of a thermophilic enzyme. Quasi-elastic neutron scattering measurements have also been performed to probe the protein motions. Compared to the free enzyme, the average height of the activation free energy barrier to local motions was found to be increased by 0.54 kcal.mol(-1) in the immobilized dihydrofolate reductase, a value that is of the same order as expected from the theoretical rate equation.
本文综述了嗜热蛋白动力学、稳定性和活性之间的关系。中子光谱已被用于测量和比较不同条件下各种嗜热和嗜温蛋白的大分子动力学。首先，对詹氏甲烷球菌的嗜热苹果酸脱氢酶和嗜温同源物兔肌肉乳酸脱氢酶进行了分子动力学分析。中子散射方法提供了对每种蛋白质整体柔韧性和结构弹性的独立测量，并且已经证明大分子动力学是热适应的分子机制之一。发现嗜热蛋白的弹性更高，从而确保两种酶在其最佳活性温度下具有相似的柔韧性。其次，已开发出中子方法来量化细胞自然拥挤环境中的平均大分子柔韧性和弹性，并在嗜冷、嗜温、嗜热和超嗜热细菌的体内测量了平均大分子运动。发现细菌中的大分子弹性随着对高温的适应而增加，而柔韧性在狭窄范围内保持，与所有处于活跃状态的细胞的生理温度无关。第三，测量了来自大肠杆菌的游离和固定化二氢叶酸还原酶中的大分子运动。固定化的嗜温酶具有更高的稳定性和更低的活性，因此其性质发生了变化，类似于嗜热酶的性质。还进行了准弹性中子散射测量以探测蛋白质运动。与游离酶相比发现，固定化二氢叶酸还原酶中局部运动的活化自由能垒的平均高度增加了0.54千卡·摩尔-1，该值与理论速率方程预期的值处于同一数量级。
