An RNA-binding complex involved in ribosome biogenesis contains a protein with homology to tRNA CCA-adding enzyme.
An RNA-binding complex involved in ribosome biogenesis contains a protein with homology to tRNA CCA-adding enzyme.
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DOI：
10.1371/journal.pbio.1001669
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发表时间：
2013-10
期刊：
影响因子：
9.8
通讯作者：
Ye K
中科院分区：
文献类型：
--
作者：
Lin J;Lu J;Feng Y;Sun M;Ye K
两个核糖体合成因子的复合物的结构和它们的核糖体结合位点的鉴定提供了对核糖体生物发生的早期阶段的见解。大量的蛋白质和小的核仁RNA与真核生物的核糖体RNA瞬时结合，以指导它们的修饰和加工以及核糖体蛋白的组装。Utp 22和Rrp 7是两个相互作用的蛋白质，没有可识别的结构域，是90 S前核糖体或小亚基加工体的组成部分，进行18 S rRNA的早期加工。在这里，我们确定的共晶体结构的Utp 22和Rrp 7复合物在1.97 μ m分辨率和NMR结构的C-末端片段的Rrp 7，这是不可见的晶体结构。结构揭示，Utp 22令人惊讶地类似于二聚体I类tRNA CCA添加酶，但具有简并活性位点，提出了tRNA和rRNA加工机制之间有趣的进化联系。Rrp 7使用异常RNA识别基序和延伸接头与Utp 22广泛结合。在酵母中通过基于结构的诱变鉴定了这两种蛋白的功能位点。我们发现，Rrp 7包含一个灵活的RNA结合的C-末端尾巴，这是必不可少的协会与前核糖体。RNA-蛋白质交联显示Rrp 7结合在18 S rRNA的中心结构域，并与两个加工H/ACA snoRNA snR 30和snR 10共享邻近区域。snR 30的缺失阻止Rrp 7稳定组装成前核糖体。我们的研究结果提供了深入了解Utp 22和Rrp 7的进化起源和功能背景。核糖体是在所有生物体中制造蛋白质的大型RNA-蛋白质复合物。大小核糖体亚基的合成是一项基本而艰巨的任务，需要真核细胞中大约200种组装因子的活性。这些因子与核糖体瞬时结合，形成一系列前核糖体颗粒。目前，我们对核糖体前体的结构和组装的了解很少。Utp 22和Rrp 7是存在于小核糖体亚基的早期前体中的两种相互作用的蛋白质。在这项研究中，我们确定的Utp 22和Rrp 7复合物的结构，通过X-射线晶体学和NMR和解剖其功能域的诱变。Utp 22的结构揭示了与tRNA CCA添加酶的意想不到的结构相似性，为深入了解Utp 22的进化起源提供了线索。utp 22显然缺乏任何酶活性，而是作为结构构建块发挥作用。Rrp 7与Utp 22广泛相关，并且似乎通过柔性RNA结合尾锚定到前核糖体。我们用RNA-蛋白质交联法鉴定了Rrp 7在前核糖体上的结合位点和邻近因子。我们的研究提供了一个详细的了解小核糖体亚基前体的结构。
The structure of a complex of two ribosome synthesis factors and identification of their ribosomal binding sites provides insights into early stages of ribosome biogenesis. A multitude of proteins and small nucleolar RNAs transiently associate with eukaryotic ribosomal RNAs to direct their modification and processing and the assembly of ribosomal proteins. Utp22 and Rrp7, two interacting proteins with no recognizable domain, are components of the 90S preribosome or the small subunit processome that conducts early processing of 18S rRNA. Here, we determine the cocrystal structure of Utp22 and Rrp7 complex at 1.97 Å resolution and the NMR structure of a C-terminal fragment of Rrp7, which is not visible in the crystal structure. The structure reveals that Utp22 surprisingly resembles a dimeric class I tRNA CCA-adding enzyme yet with degenerate active sites, raising an interesting evolutionary connection between tRNA and rRNA processing machineries. Rrp7 binds extensively to Utp22 using a deviant RNA recognition motif and an extended linker. Functional sites on the two proteins were identified by structure-based mutagenesis in yeast. We show that Rrp7 contains a flexible RNA-binding C-terminal tail that is essential for association with preribosomes. RNA–protein crosslinking shows that Rrp7 binds at the central domain of 18S rRNA and shares a neighborhood with two processing H/ACA snoRNAs snR30 and snR10. Depletion of snR30 prevents the stable assembly of Rrp7 into preribosomes. Our results provide insight into the evolutionary origin and functional context of Utp22 and Rrp7. Ribosomes are large RNA–protein complexes that manufacture proteins in all living organisms. Synthesis of large and small ribosomal subunits is a fundamental and enormous task that requires activities of approximately 200 assembly factors in eukaryotic cells. These factors transiently associate with the ribosome, forming a series of pre-ribosomal particles. We currently have a poor understanding of the structure and assembly of ribosome precursors. Utp22 and Rrp7 are two interacting proteins present in early precursors of the small ribosomal subunit. In this study, we determined the structure of the Utp22 and Rrp7 complex by X-ray crystallography and NMR and dissected their functional domains by mutagenesis. The structure of Utp22 reveals an unexpected structural similarity to the tRNA CCA-adding enzyme, providing insight into the evolutionary origin of Utp22. Utp22 apparently lacks any enzymatic activity and functions instead as a structural building block. Rrp7 associates extensively with Utp22 and appears to be anchored to pre-ribosomes via a flexible RNA-binding tail. We used RNA–protein crosslinking to identify the binding site and neighboring factor of Rrp7 on pre-ribosomes. Our study provides a detailed insight into the structure of small ribosomal subunit precursors.
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DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
14.9
作者：
Granneman, S;Gallagher, JEG;Pruijn, GJM
通讯作者：
Pruijn, GJM
影响因子：
6.8
作者：
Daubner, Gerrit M.;Clery, Antoine;Allain, Frederic H-T
通讯作者：
Allain, Frederic H-T
影响因子：
16
作者：
Grandi, P;Rybin, V;Hurt, E
通讯作者：
Hurt, E
DOI：
10.1107/s0907444909052925
发表时间：
2010-02
期刊：
Acta crystallographica. Section D, Biological crystallography
影响因子：
--
作者：
Adams PD;Afonine PV;Bunkóczi G;Chen VB;Davis IW;Echols N;Headd JJ;Hung LW;Kapral GJ;Grosse-Kunstleve RW;McCoy AJ;Moriarty NW;Oeffner R;Read RJ;Richardson DC;Richardson JS;Terwilliger TC;Zwart PH
通讯作者：
Zwart PH