环化可以稳定蛋白质的结构,如先前在单结构域蛋白质中所证明的。虽然Lys48连接的聚泛素,多结构域蛋白质,也已知在人类细胞中被环化,环化的结构效应尚不清楚。这里,我们研究了环化对环状Lys48连接的双泛素(Ub2)的结构稳定性和动力学的影响。不出所料,环化增加了Ub2的热稳定性及其对蛋白水解消化的抗性,表明环化稳定了Ub2的结构。此外,环化抑制了Ub2中的域间运动,但是环状Ub2在NMR弛豫分散实验中仍然表现出微秒构象交换。一系列长的粗粒度(CG)MD模拟显示了环化如何将Ub2的固有纳秒开闭域运动减慢到微秒。因此,CG-MD分析有助于解释意外的NMR弛豫结果,从而有助于表征环状Ub2的结构稳定性。本文受版权保护。保留所有权利。
Cyclization can stabilize the structure of proteins, as previously demonstrated in single-domain proteins. Although Lys48-linked polyubiquitin, a multi-domain protein, is also known to be cyclized in human cells, the structural effects of cyclization remain unclear. Here, we examined the impact of cyclization on the structural stability and dynamics of cyclic Lys48-linked diubiquitin (Ub2 ). As expected, cyclization increased the thermal stability of Ub2 and its resistance to proteolytic digestion, indicating that cyclization stabilized the structure of Ub2 . Furthermore, cyclization repressed the interdomain motion in Ub2 , but cyclic Ub2 still exhibited microsecond conformational exchange in NMR relaxation dispersion experiments. A series of long coarse-grained (CG) MD simulations visualized how cyclization slowed down the intrinsic nanosecond open-closed domain motion of Ub2 to microseconds. Thus, CG-MD analysis helped to explain the unexpected NMR relaxation results, thereby facilitating characterization of the structural stabilization of cyclic Ub2 .