Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, 77555, USA.
Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, 77555, USA.
Nat Commun. 2022 Jun 13;13(1):3388. doi: 10.1038/s41467-022-31129-2.
During translation initiation, initiation factor 2 (IF2) holds initiator transfer RNA (fMet-tRNA) in a specific orientation in the peptidyl (P) site of the ribosome. Upon subunit joining IF2 hydrolyzes GTP and, concomitant with inorganic phosphate (P) release, changes conformation facilitating fMet-tRNA accommodation into the P site and transition of the 70 S ribosome initiation complex (70S-IC) to an elongation-competent ribosome. The mechanism by which IF2 separates from initiator tRNA at the end of translation initiation remains elusive. Here, we report cryo-electron microscopy (cryo-EM) structures of the 70S-IC from Pseudomonas aeruginosa bound to compact IF2-GDP and initiator tRNA. Relative to GTP-bound IF2, rotation of the switch 2 α-helix in the G-domain bound to GDP unlocks a cascade of large-domain movements in IF2 that propagate to the distal tRNA-binding domain C2. The C2-domain relocates 35 angstroms away from tRNA, explaining how IF2 makes way for fMet-tRNA accommodation into the P site. Our findings provide the basis by which IF2 gates the ribosome to the elongation phase.
在翻译起始过程中，起始因子 2（IF2）将起始转移 RNA（fMet-tRNA）以特定的取向保持在核糖体的肽酰（P）位。亚基结合后，IF2 水解 GTP，伴随着无机磷酸（P）的释放，构象发生变化，促进 fMet-tRNA 进入 P 位，并使 70S 核糖体起始复合物（70S-IC）转变为具有延伸能力的核糖体。IF2 在翻译起始结束时如何从起始 tRNA 上分离仍然难以捉摸。在这里，我们报道了结合紧凑 IF2-GDP 和起始 tRNA 的铜绿假单胞菌 70S-IC 的冷冻电镜（cryo-EM）结构。与结合 GTP 的 IF2 相比，结合 GDP 的 G 结构域中的开关 2 α-螺旋的旋转解锁了 IF2 中一连串的大亚基运动，这些运动传播到远端 tRNA 结合域 C2。C2 结构域与 tRNA 分离 35 埃，解释了 IF2 如何为 fMet-tRNA 进入 P 位腾出空间。我们的发现为 IF2 将核糖体门控到延伸阶段提供了基础。
