Lu Junyan, Jiang Chenxiao, Li Xiaojing, Jiang Lizhi, Li Zengxia, Schneider-Poetsch Tilman, Liu Jianwei, Yu Kunqian, Liu Jun O, Jiang Hualiang, Luo Cheng, Dang Yongjun
Key Laboratory of Metabolism and Molecular Medicine, the Ministry of Education, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
Key Laboratory of Metabolism and Molecular Medicine, the Ministry of Education, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China.
Nucleic Acids Res. 2015 Dec 2;43(21):10157-67. doi: 10.1093/nar/gkv1033. Epub 2015 Oct 12.
Eukaryotic translation initiation factor eIF4AI, the founding member of DEAD-box helicases, undergoes ATP hydrolysis-coupled conformational changes to unwind mRNA secondary structures during translation initiation. However, the mechanism of its coupled enzymatic activities remains unclear. Here we report that a gating mechanism for Pi release controlled by the inter-domain linker of eIF4AI regulates the coupling between ATP hydrolysis and RNA unwinding. Molecular dynamic simulations and experimental results revealed that, through forming a hydrophobic core with the conserved SAT motif of the N-terminal domain and I357 from the C-terminal domain, the linker gated the release of Pi from the hydrolysis site, which avoided futile hydrolysis cycles of eIF4AI. Further mutagenesis studies suggested this linker also plays an auto-inhibitory role in the enzymatic activity of eIF4AI, which may be essential for its function during translation initiation. Overall, our results reveal a novel regulatory mechanism that controls eIF4AI-mediated mRNA unwinding and can guide further mechanistic studies on other DEAD-box helicases.
真核生物翻译起始因子eIF4AI是DEAD-box解旋酶家族的创始成员，在翻译起始过程中，它通过ATP水解偶联的构象变化来解开mRNA二级结构。然而，其偶联酶活性的机制仍不清楚。在此，我们报道由eIF4AI的结构域间连接子控制的无机磷酸（Pi）释放的门控机制调节ATP水解与RNA解旋之间的偶联。分子动力学模拟和实验结果表明，连接子通过与N端结构域保守的SAT基序和C端结构域的I357形成疏水核心，控制Pi从水解位点释放，从而避免了eIF4AI的无效水解循环。进一步的诱变研究表明，该连接子在eIF4AI的酶活性中也起自抑制作用，这可能对其在翻译起始过程中的功能至关重要。总体而言，我们的结果揭示了一种控制eIF4AI介导的mRNA解旋的新型调控机制，并可为其他DEAD-box解旋酶的进一步机制研究提供指导。
