Unstructured RNA is a substrate for tRNase Z
Unstructured RNA is a substrate for tRNase Z
复制标题
DOI：
10.1021/bi051972s
复制
发表时间：
2006-05-02
期刊：
影响因子：
2.9
通讯作者：
Nashimoto, M
中科院分区：
文献类型：
--
作者：
Shibata, HS;Minagawa, A;Nashimoto, M
TRNase Z几乎存在于所有细胞中，被认为主要作用于tRNA3‘成熟。然而，在大肠杆菌中，tRNase Z基因在正常生长条件下似乎是必不可少的，其生理作用尚不清楚。在这里，为了研究大肠杆菌tRNase Z切割前tRNAs以外的RNA的可能性，我们测试了几个非结构RNA的切割。令人惊讶的是，在体外，所有这些底物都被重组的大肠杆菌酶在多个位置非常有效地切割。枯草芽孢杆菌和黑热菌的tRNase Zs也能裂解各种非结构RNA。大肠杆菌和枯草芽孢杆菌的酶似乎有在胞苷或尿苷之前切割的趋势，而毛滴虫的酶在CCA之后除了其他切割外，不可避免地发生切割。确定最佳条件的实验表明，枯草芽孢杆菌和黑曲霉tRNase Zs对金属离子的需求不同。TRNase Z/Pre-tRNA复合体的K-d值明显高于酶/Pre-tRNA复合体。此外，来自酵母、猪和人的真核tRNase Z在多个位置切割非结构RNA，而来自嗜氧热杆菌的古生菌tRNase Z不能。
tRNase Z, which exists in almost all cells, is believed to be working primarily for tRNA 3' maturation. In Escherichia coli, however, the tRNase Z gene appears to be dispensable under normal growth conditions, and its physiological role is not clear. Here, to investigate a possibility that E. coli tRNase Z cleaves RNAs other than pre-tRNAs, we tested several unstructured RNAs for cleavage. Surprisingly, all these substrates were cleaved very efficiently at multiple sites by a recombinant E. coli enzyme in vitro. tRNase Zs from Bacillus subtilis and Thermotoga maritima also cleaved various unstructured RNAs. The E. coli and B. subtilis enzymes seem to have a tendency to cleave after cytidine or before uridine, while cleavage by the T. maritima enzyme inevitably occurred after CCA in addition to the other cleavages. Assays to determine optimal conditions indicated that metal ion requirements differ between B. subtilis and T. maritima tRNase Zs. There was no significant difference in the observed rate constant between unstructured RNA and pre-tRNA substrates, while the K-d value of a tRNase Z/unstructured RNA complex was much higher than that of an enzyme/pre-tRNA complex. Furthermore, eukaryotic tRNase Zs from yeast, pig, and human cleaved unstructured RNA at multiple sites, but an archaeal tRNase Z from Pyrobaculum aerophilum did not.