Viral Evolved Inhibition Mechanism of the RNA Dependent Protein Kinase PKR's Kinase Domain, a Structural Perspective

The protein kinase PKR activated by viral dsRNA, phosphorylates the eIF2α, which inhibit the mechanism of translation initiation. Viral evolved proteins mimicking the eIF2α block its phosphorylation and help in the viral replication. To decipher the molecular basis for the PKR’s substrate and inhibitor interaction mechanisms, we carried the molecular dynamics studies on the catalytic domain of PKR in complex with substrate eIF2α, and inhibitors TAT and K3L. The studies conducted show the altered domain movements of N lobe, which confers open and close state to the substrate-binding cavity. In addition, PKR exhibits variations in the secondary structural transition of the activation loop residues, and inter molecular contacts with the substrate and the inhibitors. Phosphorylation of the P+1 loop at the Thr-451 increases the affinity of the binding proteins exhibiting its role in the phosphorylation events. The implications of structural mechanisms uncovered will help to understand the basis of the evolution of the host-viral and the viral replication mechanisms.由病毒双链RNA（double-stranded RNA, dsRNA）激活的蛋白激酶PKR（protein kinase PKR）可对真核翻译起始因子2α（eukaryotic translation initiation factor 2α, eIF2α）进行磷酸化修饰，进而抑制翻译起始机制。病毒演化出的模拟eIF2α的蛋白质能够阻断其磷酸化过程，助力病毒复制。为解析PKR的底物与抑制剂相互作用机制的分子基础，我们针对与底物eIF2α、抑制剂TAT及K3L结合的PKR催化结构域（catalytic domain）开展了分子动力学研究。本研究结果显示，PKR的N端叶结构域（N lobe）的运动模式发生改变，使底物结合空腔（substrate-binding cavity）呈现开放与闭合两种状态。此外，PKR的激活环（activation loop）残基的二级结构转换过程存在差异，且其与底物、抑制剂之间的分子间相互作用模式亦发生变化。P+1环（P+1 loop）在苏氨酸451（Threonine-451, Thr-451）位点的磷酸化修饰可增强结合蛋白的亲和力，揭示了其在磷酸化事件中的作用。本次研究揭示的结构机制的生物学意义，将有助于解析宿主-病毒共进化以及病毒复制机制的分子基础。