Hartmann R K, Ulbrich N, Erdmann V A
Freie Universität Berlin, Institut für Biochemie, FRG.
Biochimie. 1987 Oct;69(10):1097-104. doi: 10.1016/0300-9084(87)90009-5.
We succeeded in identifying a promoter element within 200 base pairs upstream a transcriptional unit comprising only a 23S rRNA, 5S rRNA and a tRNA(gly) gene in Thermus thermophilus HB8 [1, 2]. This element shows a high degree of homology to the -35 and -10 consensus sequences for promoters described for Escherichia coli [3, 4]. The promoter activity was measured by the induction of the synthesis of functional chloramphenicol acetyltransferase in Escherichia coli. A region located at the transcriptional start, rich in guanosines and cytidines, is very similar in sequence to the one believed to be under stringent control in stable RNA and ribosomal protein genes of Escherichia coli [5]. Employing nuclease S1 protection we were able to determine the in vivo start of transcription, which was identical with the in vitro start using Escherichia coli RNA-polymerase. Furthermore we identified sequences in the region following the origin of transcription, which are homologous to sections in Escherichia coli rrn promoter-leader regions responsible for antitermination. Our finding of a promoter immediately preceding a 23S/5S rRNA operon proves a transcriptional decoupling of the 16S rRNA genes, a situation so far unprecedented among prokaryotes.
我们成功地在嗜热栖热菌HB8中一个仅包含23S rRNA、5S rRNA和一个tRNA(甘氨酸)基因的转录单元上游200个碱基对内鉴定出一个启动子元件[1,2]。该元件与大肠杆菌中所描述的启动子的-35和-10共有序列具有高度同源性[3,4]。通过在大肠杆菌中诱导功能性氯霉素乙酰转移酶的合成来测量启动子活性。位于转录起始位点的一个富含鸟苷和胞苷的区域,其序列与大肠杆菌稳定RNA和核糖体蛋白基因中被认为处于严格控制之下的区域非常相似[5]。利用核酸酶S1保护实验,我们能够确定体内转录起始位点,它与使用大肠杆菌RNA聚合酶的体外转录起始位点相同。此外,我们在转录起始位点之后的区域鉴定出了与大肠杆菌rrn启动子-前导序列区域中负责抗终止的部分同源的序列。我们在一个23S/5S rRNA操纵子之前紧邻处发现一个启动子,这证明了16S rRNA基因的转录解偶联,这种情况在原核生物中迄今为止是前所未有的。