Selection of cleavage site by mammalian tRNA 3′ processing endoribonuclease
Selection of cleavage site by mammalian tRNA 3′ processing endoribonuclease
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DOI：
10.1006/jmbi.1999.2639
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发表时间：
1999-04-09
影响因子：
5.6
通讯作者：
Kaspar, RL
中科院分区：
文献类型：
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作者：
Nashimoto, M;Tamura, M;Kaspar, RL
哺乳动物tRNA3‘加工内切核糖核酸酶(3’tRNase)通过切割识别核苷酸下游的RNA来去除前tRNAs中的3‘拖尾。尽管3‘tRNase可以识别和切割任何与另一种RNA形成类似前tRNA的复合体的靶RNA，但在某些情况下，切割发生在识别器附近的多个位置。我们使用不同的前tRNA(Arg)变异体和纯化的猪酶来研究前tRNA的哪些特征决定了切割位点。由于T茎环和受体茎加上3‘末端片段足以被3’tRNase识别，我们构建了在T茎和/或受体茎中有碱基对添加和/或缺失的变体。缺少一个和两个受体茎碱基对的前tRNA分别在鉴别子下游被切割一个和两个核苷酸以及两个和三个核苷酸。另一方面，含有额外受体茎碱基对的前tRNA变体只有在鉴别器之后才被切割。通过从T茎中删除一个碱基对，将切割位点移动到鉴别器下游的一个和两个核苷酸，但不会被T茎中的额外碱基对改变。含有8个碱基对的受体茎+6个碱基对的T茎，8个碱基对的受体茎+4个碱基对的T茎，或6个碱基对的受体茎+6个碱基对的T茎的前tRNA变体都在原始核苷酸之后被切割。通常，只有在鉴别器之后才切割受体茎和T茎中总长度超过11bp的前tRNA变体，而在鉴别器下游切割总计N个碱基(N小于12)的前tRNA变体12-N和13-N个核苷酸。变异体的切割效率取决于与真实的前tRNA相比结构变化的程度。这表明受体茎和T茎的碱基对数量对于3‘tRNase的识别和切割是重要的；(C)1999年学术出版社。
Mammalian tRNA 3' processing endoribonuclease (3' tRNase) removes 3' trailers from pre-tRNAs by cleaving the RNA immediately downstream of the discriminator nucleotide. Although 3' tRNase can recognize and cleave any target RNA that forms a pre-tRNA-like complex with another RNA, in some cases cleavage occurs at multiple sites near the discriminator. We investigated what features of pre-tRNA determine the cleavage site using various pre-tRNA(Arg) variants and purified pig enzyme. Because the T stem-loop and the acceptor stem plus a 3' trailer are sufficient for recognition by 3' tRNase, we constructed variants that had additions and/or deletions of base-pairs in the T stem and/or the acceptor stem. Pre-tRNAs lacking one and two acceptor stem base-pairs were cleaved one and two nucleotides and two and three nucleotides, respectively, downstream of the discriminator. On the other hand, pre-tRNA variants containing extra acceptor stem base-pairs were cleaved only after the discriminator. The cleavage site was shifted to one and two nucleotides downstream of the discriminator by deleting one base-pair from the T stem, but was not changed by additional base-pairs in the T stem. Pre-tRNA variants that contained an eight base-pair acceptor stem plus a six base-pair T stem, an eight base-pair acceptor stem plus a four base-pair T stem, or a six base-pair acceptor stem plus a six base-pair T stem were all cleaved after the original nucleotide. In general, pre-tRNA variants containing a total of more than 11 bp in the acceptor stem and the T stem were cleaved only after the discriminator, and pre-tRNA variants with a total of N bp (N is less than 12) were cleaved 12 - N and 13 - N nt downstream of the discriminator. Cleavage efficiency of the variants decreased depending on the degree of structural changes from the authentic pre-tRNA. This suggests that the numbers of base-pairs of both the acceptor stem and the T stem are important for recognition and cleavage by 3' tRNase; (C) 1999 Academic Press.