Pabón Germán, Amzel L Mario
Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Biophys J. 2006 Jul 15;91(2):467-72. doi: 10.1529/biophysj.106.082594. Epub 2006 Apr 21.
We have studied the unfolding by force of one of the immunoglobulin domains of the muscle protein titin using molecular dynamics simulations at 300 K. Previous studies, done at constant pulling rates, showed that under the effect of the force two strands connected to each other by six backbone H-bonds are pulled apart. No details about the mechanism of H-bond breaking were provided. Our simulation protocol "pull and wait" was designed to correspond to very slow pulling, more similar to the rates used in experiments than are the protocols used in previous computational studies. Under these conditions interstrand backbone H-bonds are not "ripped apart" by the application of the force. Instead, small elongations produced by the force weaken specific backbone H-bonds with respect to water-backbone H-bonds. These weakened bonds allow a single water molecule to make H-bonds to the CO and the NH of the same backbone H-bond while they are still bound to each other. The backbone H-bond then breaks (distance > 3.6 A), but its donor and acceptor atoms remain bound to the same water molecule. Further separation of the chains takes place when a second water molecule makes an H-bond with either the protein backbone donor or acceptor atom. Thus, the force does not directly break the main chain H-bonds: it destabilizes them in such a way that they are replaced by H-bonds to water. With this mechanism, the force necessary to break all the H-bonds required to separate the two strands will be strongly dependent on the pulling speed. Further simulations carried out at low forces but long waiting times (> or = 500 ps, < or = 10 ns) show that, given enough time, even a very small pulling force (< 400 pN) is sufficient to destabilize the interstrand H-bonds and allow them to be replaced by H-bonds to two water molecules. As expected, increasing the temperature to 350 K allows the interstrand H-bonds to break at lower forces than those required at 300 K.
我们利用300K下的分子动力学模拟研究了肌肉蛋白肌联蛋白的一个免疫球蛋白结构域在力作用下的展开过程。先前在恒定拉伸速率下进行的研究表明，在力的作用下，通过六个主链氢键相互连接的两条链会被拉开。但未提供有关氢键断裂机制的详细信息。我们设计的“拉伸并等待”模拟方案旨在对应非常缓慢的拉伸，与先前计算研究中使用的方案相比，更类似于实验中使用的速率。在这些条件下，链间主链氢键不会因力的作用而“被撕开”。相反，力产生的小伸长会使特定的主链氢键相对于水 - 主链氢键变弱。这些变弱的键允许单个水分子在它们仍然相互结合时与同一主链氢键的羰基（CO）和亚氨基（NH）形成氢键。然后主链氢键断裂（距离> 3.6埃），但其供体和受体原子仍与同一个水分子结合。当第二个水分子与蛋白质主链供体或受体原子形成氢键时，链会进一步分离。因此，力不会直接断裂主链氢键：它以使它们被与水的氢键取代的方式使其不稳定。通过这种机制，分离两条链所需的所有氢键断裂所需的力将强烈依赖于拉伸速度。在低力但长等待时间（≥500皮秒，≤10纳秒）下进行的进一步模拟表明，给予足够的时间，即使非常小的拉伸力（<400皮牛顿）也足以使链间氢键不稳定，并使其被与两个水分子的氢键取代。正如预期的那样，将温度升高到350K会使链间氢键在比300K所需的力更低的力下断裂。
