Reconstitution of full-round uridine-deletion RNA editing with three recombinant proteins
Reconstitution of full-round uridine-deletion RNA editing with three recombinant proteins
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DOI：
10.1073/pnas.0604476103
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发表时间：
2006-09-19
影响因子：
11.1
通讯作者：
Simpson, Larry
中科院分区：
文献类型：
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作者：
Kang, Xuedong;Gao, Guanghan;Simpson, Larry
锥虫线粒体中的尿苷（U）-插入/缺失RNA编辑涉及通过部分互补的指导RNA的退火确定的特定位点处的预编辑mRNA的初始切割。已经提出了两种含有RNase III的核心编辑复合物（L-复合物）蛋白MP 90（KREPB 1）和MP 61（KREPB 3）分别参与U-缺失和U-插入编辑，但是这些推定的酶尚未被表征或以活性形式表达。重组蛋白MP 90从布氏锥虫和利什曼原虫主要分别在昆虫细胞和tarentolae利什曼原虫胞质溶胶中表达。这些蛋白质在特异性切割模型U-缺失位点而不是U-插入位点中是有活性的。RNase III基序的缺失或突变消除了这种活性。用重组MP 90和来自L. major和重组RNA编辑RNA连接酶1。tarentolae MP 90被命名为REN 1，用于RNA编辑核酸酶1。
Uridine (U)-insertion/deletion RNA editing in trypanosome mitochondria involves an initial cleavage of the preedited mRNA at specific sites determined by the annealing of partially complementary guide RNAs. An involvement of two RNase Ill-containing core editing complex (L-complex) proteins, MP90 (KREPB1) and MP61 (KREPB3) in, respectively, U-deletion and U-insertion editing, has been suggested, but these putative enzymes have not been characterized or expressed in active form. Recombinant MP90 proteins from Trypanosoma brucei and Leishmania major were expressed in insect cells and cytosol of Leishmania tarentolae, respectively. These proteins were active in specifically cleaving a model U-deletion site and not a U-insertion site. Deletion or mutation of the RNase III motif abolished this activity. Full-round guide RNA (gRNA)-mediated in vitro U-deletion editing was reconstituted by a mixture of recombinant MP90 and recombinant RNA editing exonuclease I from L. major, and recombinant RNA editing RNA ligase 1 from L. tarentolae. MP90 is designated REN1, for RNA-editing nuclease 1.