Processing of multiple-intron-containing pretRNA
Processing of multiple-intron-containing pretRNA
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DOI：
10.1073/pnas.0911658106
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发表时间：
2009-12
期刊：
影响因子：
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通讯作者：
G. Tocchini-Valentini;P. Fruscoloni;G. Tocchini-Valentini
中科院分区：
文献类型：
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作者：
G. Tocchini-Valentini;P. Fruscoloni;G. Tocchini-Valentini
计算研究预测在某些crenarchaeal tRNA基因中同时存在两个甚至三个内含子。在这些含有多个内含子的pretRNA中，内含子一个嵌套在另一个内部，pretRNA折叠成一种构象，预计只有在拼接其他内含子之后才能拼接最后一个内含子。一组操作，每个操作由两个裂解和一个结扎组成，因此需要按顺序进行。预计含有多个内含子的pretrna会折叠，形成凸起-螺旋-凸起（BHB）和BHB样基序。tRNA剪接内切酶应该能识别这些基序。为了验证这一假设，我们使用了来自jannaschii甲烷醛球菌（METJA）的同四聚体酶和来自solfataricus （SULSO）的异四聚体酶。根据我们之前的研究，METJA酶只能切割BHB结构基序，而SULSO酶还可以切割不同的底物结构，如BHL结构。我们在这里展示了可以在体外观察到含有多个内含子的pretRNA的加工。
Computational studies predict the simultaneous presence of two and even three introns in certain crenarchaeal tRNA genes. In these multiple-intron-containing pretRNAs, the introns are nested one inside the other and the pretRNA folds into a conformation that is anticipated to allow splicing of the last intron only after splicing the others. A set of operations, each consisting of two cleavages and one ligation, therefore needs to be carried out sequentially. PretRNAs containing multiple introns are predicted to fold, forming bulge–helix–bulge (BHB) and BHB-like motifs. The tRNA splicing endonuclease should recognize these motifs. To test this hypothetical scenario, we used the homotetrameric enzyme from Methanocaldococcus jannaschii (METJA) and the heterotetrameric enzyme from Sulfolobus solfataricus (SULSO). On the basis of our previous studies, the METJA enzyme should cleave only the BHB structure motif, while the SULSO enzyme can in addition cleave variant substrate structures, like the bulge-helix-loop (BHL). We show here that the processing of multiple-intron-containing pretRNA can be observed in vitro.