A TRANSFER-RNA-LIKE STRUCTURE IS PRESENT IN 10SA RNA, A SMALL STABLE RNA FROM ESCHERICHIA-COLI
A TRANSFER-RNA-LIKE STRUCTURE IS PRESENT IN 10SA RNA, A SMALL STABLE RNA FROM ESCHERICHIA-COLI
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DOI：
10.1073/pnas.91.20.9223
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发表时间：
1994-09-27
影响因子：
11.1
通讯作者：
INOKUCHI, H
中科院分区：
文献类型：
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作者：
KOMINE, Y;KITABATAKE, M;INOKUCHI, H
我们确定10Sa RNA（大肠杆菌中发现的小稳定RNA之一）具有一个有趣的结构特征：10Sa RNA的5'端和3'端可以排列成相当于大肠杆菌丙氨酸tRNA的半分子（受体茎和TFC茎环）的结构。对从具有温度敏感 RNase P 功能的细菌突变体中提取的 10Sa RNA 进行引物延伸分析表明，10Sa RNA 的前体（pre-10Sa RNA）在体内折叠成类似前 tRNA 的结构，从而可以被 RNase P 切割，生成成熟 10Sa RNA 的 5' 端。纯化的10Sa RNA可以在体外荷上丙氨酸。编码 10Sa RNA (ssrA) 的基因遭到破坏会导致细胞生长速率降低（在 45 摄氏度时尤其明显），并且会导致半固体琼脂上的运动性降低。缺失菌株 (Delta ssrA) 的这些表型特征使我们能够研究 10Sa RNA 体内某些突变的影响，尽管 10Sa RNA 的确切功能仍不清楚。当10Sa RNA中的G U对（G3-U357）（可能相当于丙氨酸tRNA的决定性G.U对）改变为G A或G.C对时，补充Delta ssrA菌株表型突变的能力就丧失了。此外，通过在多拷贝质粒上引入编码丙氨酰-tRNA合成酶（alaS）的基因，可以克服由于G.A对取代决定簇碱基而导致的无法补充突变表型的问题。证据表明，所提出的 10Sa RNA 结构特征确实在体内表现出来。
We have determined that 10Sa RNA (one of the small stable RNAs found in Escherichia coli) has an interesting structural feature: the 5' end and the 3' end of 10Sa RNA can be arranged in a structure that is equivalent to a half-molecule (acceptor stem and TFC stem-loop) of alanine tRNA of E. coli. Primer-extension analysis of 10Sa RNA extracted from a bacterial mutant with temperature-sensitive RNase P function revealed that the precursor to 10Sa RNA (pre-10Sa RNA) is folded into a pre-tRNA-like structure in vivo such that it can be cleaved by RNase P to generate the 5' end of the mature 10Sa RNA. The purified 10Sa RNA can be charged with alanine in vitro. Disruption of the gene encoding 10Sa RNA (ssrA) caused a reduction in the rate of cell growth, which was especially apparent at 45 degrees C, and a reduction in motility on semisolid agar. These phenotypic characteristics of the deletion strain (Delta ssrA) allowed us to investigate the effects of some mutations in 10Sa RNA in vivo, although the exact function of 10Sa RNA still remains unclear. When the G U pair (G3-U357) in 10Sa RNA, which may be equivalent to the determinant G.U pair of alanine tRNA, was changed to a G A or G.C pair, the ability to complement the phenotypic mutations of the Delta ssrA strain was lost. Furthermore, this inability to complement the mutant phenotypes that was caused by the substitution of the determinant bases by a G.A pair could be overcome by the introduction of a gene encoding alanyl-tRNA synthetase (alaS) on a multicopy plasmid. The evidence suggests that the proposed structural features of 10Sa RNA are indeed manifested in vivo.