Functional requirement for symmetric assembly of archaeal box C/D small ribonucleoprotein particles
Functional requirement for symmetric assembly of archaeal box C/D small ribonucleoprotein particles
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DOI：
10.1016/j.jmb.2003.08.012
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发表时间：
2003-10-17
影响因子：
5.6
通讯作者：
Li, H
中科院分区：
文献类型：
--
作者：
Rashid, R;Aittaleb, M;Li, H
盒C/D小核糖核蛋白颗粒(SRNPs)是真核生物中小核仁核糖核蛋白颗粒(SnoRNPs)的古老同源物，负责核糖体和转运体RNA的2‘-O位点特异性甲基化。盒C/D rRNPs的功能是由三种核心蛋白在盒C/D RNA周围逐步组装而成，包括纤维蛋白原、Nop5p和L7Ae。在所有的盒C/D RNA中，最明显的结构特征是存在两个保守的盒C/D基序，通常伴随着一个单独的，有时是两个反义元件，位于D或D‘盒的上游。尽管反义元件的这种不对称分布，盒C/D基序的两部分特征似乎与最近报道的纤维蛋白与Nop5p之间的核心蛋白复合体的三维结构令人满意地一致。本研究探讨了盒C/D RNA和纤维蛋白-Nop5p复合体中对称特征的功能含义。利用定点突变技术获得了缺失两个box C/D基序之一的box C/D RNA和缺乏自结合能力的突变型纤维蛋白-Nop5p复合体。通过凝胶迁移率改变、分析超速离心法和体外催化研究来评估突变组分组装和指导甲基转移反应的能力。本文的结果表明，虽然box C/D sRNP能够不对称组装，但box C/D RNA和纤维蛋白-Nop5p复合体的对称性是有效催化所必需的。这些发现强调了功能组装在甲基转移反应中的重要性。(C)2003爱思唯尔有限公司。保留所有权利。
Box C/D small ribonucleoprotein particles (sRNPs) are archaeal homologs of small nucleolar ribonucleoprotein particles (snoRNPs) in eukaryotes that are responsible for site specific 2'-O-methylation of ribosomal and transfer RNAs. The function of box C/D sRNPs is characterized by step-wise assembly of three core proteins around a box C/D RNA that include fibrillarin, Nop5p, and L7Ae. The most distinct structural feature in all box C/D RNAs is the presence of two conserved box C/D motifs accompanied by often a single, and sometimes two, antisense elements located immediately upstream of either the D or D' box. Despite this asymmetric distribution of antisense elements, the bipartite feature of the box C/D motifs appears to be in pleasing agreement with a recently reported three-dimensional structure of the core protein complex between fibrillarin and Nop5p. This investigates functional implications of the symmetric features both in box C/D RNAs and in the fibrillarin-Nop5p complex. Site-directed mutagenesis was employed to generate box C/D RNAs lacking one of the two box C/D motifs and a mutant fibrillarin-Nop5p complex deficient in self-association. The ability of the mutated components to assemble and to direct methyl transfer reactions was assessed by gel mobility-shift, analytical ultracentrifugation, and in vitro catalysis studies. The results presented here suggest that, while a box C/D sRNP is capable of asymmetrical assembly, the symmetries in both the box C/D RNA and in the fibrillarin-Nop5p complex are required for efficient catalysis. These findings underscore the importance of functional assembly in methyl transfer reactions. (C) 2003 Elsevier Ltd. All rights reserved.