X-ray structures of U2 snRNA - Branchpoint duplexes containing conserved pseudouridines
X-ray structures of U2 snRNA - Branchpoint duplexes containing conserved pseudouridines
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DOI：
10.1021/bi7022392
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发表时间：
2008-05-20
期刊：
影响因子：
2.9
通讯作者：
Kielkopf, Clara L.
中科院分区：
文献类型：
--
作者：
Lin, Yuan;Kielkopf, Clara L.
U2 小核 (sn)RNA 的假尿苷修饰区域与内含子分支点序列退火，并定位凸出的腺苷，以在前 mRNA 剪接的第一个化学步骤中充当亲核试剂。我们已经确定了包含假尿苷化 U2 snRNA 和分支点共有序列的 RNA 寡核苷酸的三种 X 射线结构。预期的腺苷分支点在包含哺乳动物共有序列变体的 1.65 埃分辨率结构和包含缩短的酿酒酵母共有序列的 2. 10 埃分辨率结构中是螺旋外的。与预期分支点相邻的腺苷在第三个结构中是螺旋外的，该结构包含分辨率为 1.57 埃的完整酵母共有序列。 U2 snRNA 假尿苷介导的水合和碱基堆积相互作用与未配对腺苷的身份相关。预期的腺苷凸起与堆叠良好的假尿苷相关，假尿苷通过有序的水分子与相邻的核苷酸连接。相反，相邻腺苷的凸起会改变碱基堆积并破坏假尿苷的水介导的相互作用。这些结构差异可能有助于假尿苷修饰促进分支位点腺苷的凸出构象并增强 snRNA 的催化作用。此外，在非常规凸出的腺苷附近鉴定出碘化物结合位点，并且哺乳动物共有序列变体的结构提供了水合镁离子以类似方式与第二组内含子的二价阳离子结合位点结合的高分辨率视图。
A pseudouridine-modified region of the U2 small nuclear (sn)RNA anneals with the intronic branchpoint sequence and positions a bulged adenosine to serve as the nucleophile in the first chemical step of pre-mRNA splicing. We have determined three X-ray structures of RNA oligonucleotides containing the pseudouridylated U2 snRNA and the branchpoint consensus sequences. The expected adenosine branchpoint is extrahelical in a 1.65 angstrom resolution structure containing the mammalian consensus sequence variant and in a 2. 10 angstrom resolution structure containing a shortened Saccharomyces cerevisiae consensus sequence. The adenosine adjacent to the expected branchpoint is extrahelical in a third structure, which contains the intact yeast consensus sequence at 1.57 angstrom resolution. The hydration and base stacking interactions mediated by the U2 snRNA pseudouridines correlate with the identity of the unpaired adenosine. The expected adenosine bulge is associated with a well-stacked pseudouridine, which is linked via an ordered water molecule to a neighboring nucleotide. In contrast, the bulge of the adjacent adenosine shifts the base stacking and disrupts the water-mediated interactions of the pseudouridine. These structural differences may contribute to the ability of the pseudouridine modification to promote the bulged conformation of the branch site adenosine and to enhance catalysis by snRNAs. Furthermore, iodide binding sites are identified adjacent to the unconventional bulged adenosine, and the structure of the mammalian consensus sequence variant provides a high-resolution view of a hydrated magnesium ion bound in a similar manner to a divalent cation binding site of the group II intron.