From the Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125 and.
J Biol Chem. 2013 Dec 20;288(51):36385-97. doi: 10.1074/jbc.M113.513614. Epub 2013 Oct 22.
The signal recognition particle (SRP) RNA is a universally conserved and essential component of the SRP that mediates the co-translational targeting of proteins to the correct cellular membrane. During the targeting reaction, two functional ends in the SRP RNA mediate distinct functions. Whereas the RNA tetraloop facilitates initial assembly of two GTPases between the SRP and SRP receptor, this GTPase complex subsequently relocalizes ∼100 Å to the 5',3'-distal end of the RNA, a conformation crucial for GTPase activation and cargo handover. Here we combined biochemical, single molecule, and NMR studies to investigate the molecular mechanism of this large scale conformational change. We show that two independent sites contribute to the interaction of the GTPase complex with the SRP RNA distal end. Loop E plays a crucial role in the precise positioning of the GTPase complex on these two sites by inducing a defined bend in the RNA helix and thus generating a preorganized recognition surface. GTPase docking can be uncoupled from its subsequent activation, which is mediated by conserved bases in the next internal loop. These results, combined with recent structural work, elucidate how the SRP RNA induces GTPase relocalization and activation at the end of the protein targeting reaction.
信号识别颗粒 (SRP) RNA 是 SRP 中普遍存在且必不可少的组成部分，它介导蛋白质在共翻译过程中靶向到正确的细胞膜。在靶向反应过程中，SRP RNA 的两个功能末端介导不同的功能。虽然 RNA 四螺旋体促进了 SRP 和 SRP 受体之间两种 GTPase 的初始组装，但该 GTPase 复合物随后会重新定位到 RNA 的 5'、3'-远端，这种构象对于 GTPase 的激活和货物交接至关重要。在这里，我们结合生化、单分子和 NMR 研究来研究这种大规模构象变化的分子机制。我们表明，两个独立的位点有助于 GTPase 复合物与 SRP RNA 远端的相互作用。Loop E 通过在 RNA 螺旋上诱导特定的弯曲，从而产生预组织的识别表面，在这两个位点上对 GTPase 复合物的精确定位起着至关重要的作用。GTPase 的对接可以与其随后的激活解耦，而后者是由内部环中的保守碱基介导的。这些结果，结合最近的结构研究，阐明了 SRP RNA 如何在蛋白质靶向反应结束时诱导 GTPase 的重新定位和激活。
