Purified box C/D snoRNPs are able to reproduce site-specific 2'-O-methylation of target RNA in vitro
Purified box C/D snoRNPs are able to reproduce site-specific 2'-O-methylation of target RNA in vitro
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DOI：
10.1128/mcb.22.19.6663-6668.2002
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发表时间：
2002-10-01
影响因子：
5.3
通讯作者：
Bozzoni, I
中科院分区：
文献类型：
--
作者：
Galardi, S;Fatica, A;Bozzoni, I
核仁小RNA（snoRNA）与位于核仁的核糖核蛋白颗粒（snoRNP）相关。盒C/D家族的大多数成员在指导底物RNA的位点特异性2 '-O-甲基化中起作用。尽管靶核苷酸的选择需要反义元件和snoRNA的保守盒D或D'，但甲基转移酶活性应该存在于蛋白质组分之一中。通过对snoRNP特异性因子进行蛋白质标记，我们从酿酒酵母中纯化了同源盒C/D snoRNP。质谱分析表明，Nop 1 p，Nop 58 p，Nop 56 p和Snu 13 p作为颗粒的组成部分的存在。我们表明，纯化的snoRNP能够再现靶RNA上的位点特异性甲基化模式，并且预测的Nop 1 p的S-腺苷-L-甲硫氨酸结合区负责催化活性。
Small nucleolar RNAs (snoRNAs) are associated in ribonucleoprotein particles localized to the nucleolus (snoRNPs). Most of the members of the box C/D family function in directing site-specific 2'-O-methylation of substrate RNAs. Although the selection of the target nucleotide requires the antisense element and the conserved box D or D' of the snoRNA, the methyltransferase activity is supposed to reside in one of the protein components. Through protein tagging of a snoRNP-specific factor, we purified to homogeneity box C/D snoRNPs from the yeast Saccharomyces cerevisiae. Mass spectrometric analysis demonstrated the presence of Nop1p, Nop58p, Nop56p, and Snu13p as integral components of the particle. We show that purified snoRNPs are able to reproduce the site-specific methylation pattern on target RNA and that the predicted S-adenosyl-L-methionine-binding region of Nop1p is responsible for the catalytic activity.