Department of Chemistry, Lensfield Road, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
J Am Chem Soc. 2011 Jan 26;133(3):513-26. doi: 10.1021/ja107863z. Epub 2011 Jan 4.
Identifying and understanding the differences between protein folding in bulk solution and in the cell is a crucial challenge facing biology. Using Langevin dynamics, we have simulated intact ribosomes containing five different nascent chains arrested at different stages of their synthesis such that each nascent chain can fold and unfold at or near the exit tunnel vestibule. We find that the native state is destabilized close to the ribosome surface due to an increase in unfolded state entropy and a decrease in native state entropy; the former arises because the unfolded ensemble tends to behave as an expanded random coil near the ribosome and a semicompact globule in bulk solution. In addition, the unfolded ensemble of the nascent chain adopts a highly anisotropic shape near the ribosome surface and the cooperativity of the folding-unfolding transition is decreased due to the appearance of partially folded structures that are not populated in bulk solution. The results show, in light of these effects, that with increasing nascent chain length folding rates increase in a linear manner and unfolding rates decrease, with larger and topologically more complex folds being the most highly perturbed by the ribosome. Analysis of folding trajectories, initiated by temperature quench, reveals the transition state ensemble is driven toward compaction and greater native-like structure by interactions with the ribosome surface and exit vestibule. Furthermore, the diversity of folding pathways decreases and the probability increases of initiating folding via the N-terminus on the ribosome. We show that all of these findings are equally applicable to the situation in which protein folding occurs during continuous (non-arrested) translation provided that the time scales of folding and unfolding are much faster than the time scale of monomer addition to the growing nascent chain, which results in a quasi-equilibrium process. These substantial ribosome-induced perturbations to almost all aspects of protein folding indicate that folding scenarios that are distinct from those of bulk solution can occur on the ribosome.
确定并理解在体溶液和细胞中蛋白质折叠的差异是生物学面临的一个关键挑战。我们使用 Langevin 动力学模拟了完整的核糖体，其中包含五个不同的新生链，这些新生链在其合成的不同阶段被捕获，使得每个新生链都可以在出口隧道前庭处折叠和展开。我们发现，由于未折叠状态的熵增加和天然状态的熵减少，天然状态在靠近核糖体表面时被破坏；前者是因为在核糖体附近，未折叠的集合倾向于表现为展开的随机线圈，而在体溶液中则表现为半紧凑的球体。此外，新生链的未折叠集合在核糖体表面附近采用高度各向异性的形状，并且由于出现了在体溶液中未出现的部分折叠结构，折叠-展开转变的协同性降低。结果表明，考虑到这些效应，随着新生链长度的增加，折叠速率以线性方式增加，展开速率降低，并且较大和拓扑上更复杂的折叠受到核糖体的干扰最大。通过温度淬火引发的折叠轨迹分析表明，过渡态集合通过与核糖体表面和出口前庭的相互作用被推向紧凑化和更类似于天然的结构。此外，折叠途径的多样性减少，并且通过核糖体上的 N 端起始折叠的概率增加。我们表明，所有这些发现对于在连续（非捕获）翻译过程中发生蛋白质折叠的情况同样适用，前提是折叠和展开的时间尺度比单体添加到生长中的新生链的时间尺度快得多，这导致准平衡过程。核糖体对蛋白质折叠几乎所有方面的这些实质性干扰表明，在核糖体上可以发生与体溶液不同的折叠情况。
