Protein-induced conformational changes of RNA during the assembly of human signal recognition particle
Protein-induced conformational changes of RNA during the assembly of human signal recognition particle
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DOI：
10.1016/j.jmb.2006.12.056
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发表时间：
2007-03-16
影响因子：
5.6
通讯作者：
Nagai, Kiyoshi
中科院分区：
文献类型：
--
作者：
Menichelli, Elena;Isel, Catherine;Nagai, Kiyoshi
人类信号识别颗粒 (SRP) 是一种大型 RNA-蛋白质复合物，可将分泌蛋白和膜蛋白靶向内质网膜。 SRP 的 S 结构域由大约一半的 7SL RNA 和四种蛋白质（SRP19、SRP54 和 SRP68/72 异二聚体）组成。为了了解蛋白质的结合如何诱导 RNA 的构象变化并影响其他蛋白质亚基的后续结合，我们对所有 S 结构域组装中间体进行了化学和酶促探测。乙基化干扰实验表明，对于 SRP68/72 的结合至关重要的螺旋 5、6 和 7 中的磷酸基团都位于 RNA 的同一面上。羟基自由基足迹和硫酸二甲酯 (DMS) 修饰表明 SRP68/72 使螺旋 6 和 8 的下部更接近。 SRP68/72 结合还可以保护 SRP54 结合位点（螺旋 8 不对称环）免受化学修饰和 RNase 裂解，而在 SRP19 和 SRP68/72 都存在的情况下，螺旋 8 不对称环的长链变得易于化学和酶探针接近。这些结果表明，在SRP19-SRP54M-RNA复合物的晶体结构中观察到的RNA平台在SRP68/72和SRP19存在的情况下已经存在。因此，SRP68/72 与 SRP19 一起将 7SL RNA 重新排列为 SRP54 结合能力状态。 (c) 2007 Elsevier Ltd. 保留所有权利。
The human signal recognition particle (SRP) is a large RNA-protein complex that targets secretory and membrane proteins to the endoplasmic reticulum membrane. The S domain of SRP is composed of roughly half of the 7SL RNA and four proteins (SRP19, SRP54, and the SRP68/72 heterodimer). In order to understand how the binding of proteins induces conformational changes of RNA and affects subsequent binding of other protein subunits, we have performed chemical and enzymatic probing of all S domain assembly intermediates. Ethylation interference experiments show that phosphate groups in helices 5, 6 and 7 that are essential for the binding of SRP68/72 are all on the same face of the RNA. Hydroxyl radical footprinting and dimethylsulphate (DMS) modifications show that SRP68/72 brings the lower part of helices 6 and 8 closer. SRP68/72 binding also protects the SRP54 binding site (helix 8 asymmetric loop) from chemical modification and RNase cleavage, whereas, in the presence of both SRP19 and SRP68/72, the long strand of helix 8 asymmetric loop becomes readily accessible to chemical and enzymatic probes. These results indicate that the RNA platform observed in the crystal structure of the SRP19-SRP54M-RNA complex already exists in the presence of SRP68/72 and SRP19. Therefore, SRP68/72, together with SRP19, rearranges the 7SL RNA in an SRP54 binding competent state. (c) 2007 Elsevier Ltd. All rights reserved.