Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria
Structure of the core editing complex (L-complex) involved in uridine insertion/deletion RNA editing in trypanosomatid mitochondria
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DOI：
10.1073/pnas.0901754106
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发表时间：
2009-07-28
影响因子：
11.1
通讯作者：
Simpson, Larry
中科院分区：
文献类型：
--
作者：
Li, Feng;Ge, Peng;Simpson, Larry
尿苷插入/缺失RNA编辑是一种独特的转录后RNA加工形式，发生在着丝质体原生生物的线粒体中。我们对通过串联亲和纯化程序（TAP）从利什曼原虫（Leishmania tarentolae）线粒体分离的核心编辑复合体或“L（连接酶）复合体”进行了3D结构分析。纯化后的材料在20-25S沉淀，在1 MDa的蓝色天然凝胶中迁移，并表现出预裂解和全周期grna介导的u插入和u缺失的体外活性。通过电子断层扫描分析纯化的l -配合物以确定异质性的程度。层析成像中单个颗粒的三维结构比较显示，大多数复合物具有类似的细长三角形形状。一个独立的单粒子重建，使用一个无特征的高斯球作为初始模型，收敛到一个类似的三角形结构。另一个单粒子重建，使用平均层析成像结构作为初始模型，产生了类似的结构。用REL1特异性IgG进行修饰，将REL1连接酶定位在模型顶端的基部。这种结构对于编辑反应的详细分析应该是有用的。
Uridine insertion/deletion RNA editing is a unique form of posttranscriptional RNA processing that occurs in mitochondria of kinetoplastid protists. We have carried out 3D structural analyses of the core editing complex or "L (ligase)-complex'' from Leishmania tarentolae mitochondria isolated by the tandem affinity purification procedure (TAP). The purified material, sedimented at 20-25S, migrated in a blue native gel at 1 MDa and exhibited both precleaved and full-cycle gRNA-mediated U-insertion and U-deletion in vitro activities. The purified L-complex was analyzed by electron tomography to determine the extent of heterogeneity. Three-dimensional structural comparisons of individual particles in the tomograms revealed that a majority of the complexes have a similar shape of a slender triangle. An independent single-particle reconstruction, using a featureless Gaussian ball as the initial model, converged to a similar triangular structure. Another single-particle reconstruction, using the averaged tomography structure as the initial model, yielded a similar structure. The REL1 ligase was localized on the model to the base of the apex by decoration with REL1-specific IgG. This structure should prove useful for a detailed analysis of the editing reaction.