Hartmann R K, Erdmann V A
Institut für Biochemie, Freie Universität Berlin, FRG.
Nucleic Acids Res. 1991 Nov 11;19(21):5957-64. doi: 10.1093/nar/19.21.5957.
The gene for the RNA subunit of ribonuclease P from the extreme thermophilic eubacterium T. thermophilus HB8 was cloned using oligonucleotide probes complementary to conserved regions of RNase P RNA subunits from proteobacteria. The monocistronic gene and its flanking regions were sequenced. The gene is enclosed by a promoter and a rho-independent terminator. Nuclease S1 protection analyses showed that the primary transcript is identical with the mature RNA, i.e. no processing events are involved. The stem and loop structure of the terminator remains part of the mature molecule. In vitro transcription of the cloned gene with purified RNA polymerase from T. thermophilus yields the same RNA product as in vivo, indicating that no other components except RNA polymerase are involved in the synthesis of the RNA. RNase P RNA from T. thermophilus cleaved a pre-tRNA(Tyr) from E. coli with highest efficiency between 55 degrees C and 65 degrees C. The T. thermophilus RNA, which has a G-C content of 86% in helical regions, displays several structural idiosyncrasies, although its secondary structure is similar to that of proteobacteria. Numerous invariable nucleotides in the structural core of eubacterial RNase P RNAs are also conserved in the RNA from the extreme thermophilic eubacterium.
利用与变形菌门核糖核酸酶P RNA亚基保守区域互补的寡核苷酸探针，克隆了嗜热栖热菌HB8核糖核酸酶P的RNA亚基基因。对单顺反子基因及其侧翼区域进行了测序。该基因由一个启动子和一个不依赖ρ因子的终止子包围。核酸酶S1保护分析表明，初级转录本与成熟RNA相同，即不涉及加工事件。终止子的茎环结构仍是成熟分子的一部分。用嗜热栖热菌纯化的RNA聚合酶对克隆基因进行体外转录，产生的RNA产物与体内相同，表明RNA合成过程中除RNA聚合酶外不涉及其他成分。嗜热栖热菌的核糖核酸酶P RNA在55℃至65℃之间能最有效地切割大肠杆菌的前体tRNA（Tyr）。嗜热栖热菌的RNA在螺旋区域的G-C含量为86%，虽然其二级结构与变形菌门的相似，但也显示出一些结构特性。真细菌核糖核酸酶P RNA结构核心中的许多不变核苷酸在嗜热真细菌的RNA中也保守。
