Biochemical identification of Argonaute 2 as the sole protein required for RNA-induced silencing complex activity
Biochemical identification of Argonaute 2 as the sole protein required for RNA-induced silencing complex activity
复制标题
DOI：
10.1073/pnas.0405913101
复制
发表时间：
2004-10-05
影响因子：
11.1
通讯作者：
Wang, XD
中科院分区：
文献类型：
--
作者：
Rand, TA;Ginalski, K;Wang, XD
RNA 干扰是由小型双链 RNA 诱导沉默复合物 (RISC) 进行的。 RISC 结合的小 RNA 引导 RISC 复合体识别和切割具有互补序列的 mRNA。构成 RISC 复合体和切割 mRNA 的蛋白质尚未明确定义。在这里，我们报告了果蝇 Schnieder 2 细胞提取物中 RISC 活性的生化纯化，使其达到均质。 Argonaute 2 (Ago-2) 是纯化的功能性 RISC 中存在的唯一蛋白质成分。通过使用序列图谱分析与预测蛋白质二级结构相结合的生物信息学方法，我们发现了Ago-2的PIWI结构域与核酸内切酶V之间的同源性，并鉴定了Ago-2的PIWI结构域内潜在的活性位点氨基酸残基。
RNA interference is carried out by the small double-stranded RNA-induced silencing complex (RISC). The RISC-bound small RNA guides the RISC complex to identify and cleave mRNAs with complementary sequences. The proteins that make up the RISC complex and cleave mRNA have not been unequivocally defined. Here, we report the biochemical purification of RISC activity to homogeneity from Drosophila Schnieder 2 cell extracts. Argonaute 2 (Ago-2) is the sole protein component present in the purified, functional RISC. By using a bioinformatics method that combines sequence-profile analysis with predicted protein secondary structure, we found homology between the PIWI domain of Ago-2 and endonuclease V and identified potential active-site amino acid residues within the PIWI domain of Ago-2.