de Zamaroczy M, Delorme F, Elmerich C
Unité de Physiologie Cellulaire and CNRS URA 1300, Département des Biotechnologies, Institut Pasteur, Paris, France.
Mol Gen Genet. 1990 Dec;224(3):421-30. doi: 10.1007/BF00262437.
The complete nucleotide sequence of the open reading frame (ORF) located upstream of the glnA structural gene for glutamine synthetase (GS) in Azospirillum brasilense Sp7 was determined. This ORF, which codes for a 12 kDa protein, was identified as glnB, the structural gene for the PII protein, a component of the adenylylation cascade involved in the regulation of GS activity in some gram-negative bacteria. Transcription analysis and mRNA mapping of glnB and glnA of A. brasilense was performed with bacteria grown under different physiological conditions. The glnA gene can be transcribed either as a glnB-A mRNA of 2.4 kb or as a glnA mRNA of 1.5 kb. Differential expression of the two mRNAs was found to depend on the nitrogen source. The glnB-A mRNA was the major transcript under nitrogen fixation conditions, while the synthesis of the glnA mRNA was almost completely abolished. The glnA mRNA was predominantly produced in NH4(+)-containing medium. Transcription start site analysis revealed the presence of three different types of nitrogen-regulated promoters. GlnB-A mRNA was transcribed selectively from tandem promoters. One of them is similar to the NtrA-dependent promoter and the other to the Escherichia coli sigma 70 promoter. The synthesis of glnA mRNA was regulated by a promoter, which was repressed (or non-activated) only under conditions of nitrogen fixation, when moleuclar nitrogen was the sole nitrogen source. The transcriptional initiation site in front of glnA is not preceded by a canonical E. coli sigma 70 promoter. A sequence reminiscent of the NtrA-dependent promoter consensus, except for a fundamental mismatch, was found at positions -33 to -21. This sequence overlapped a putative "weak" NtrC-binding site, similar to those identified in enteric bacteria. From these results, it is postulated that glnA mRNA is controlled by a novel type of nitrogen-regulated promoter.
测定了巴西固氮螺菌Sp7中谷氨酰胺合成酶（GS）的glnA结构基因上游开放阅读框（ORF）的完整核苷酸序列。该ORF编码一种12 kDa的蛋白质，被鉴定为glnB，即PII蛋白的结构基因，PII蛋白是某些革兰氏阴性细菌中参与GS活性调节的腺苷酸化级联反应的一个组分。利用在不同生理条件下生长的细菌对巴西固氮螺菌的glnB和glnA进行了转录分析和mRNA定位。glnA基因可以转录为2.4 kb的glnB - A mRNA，也可以转录为1.5 kb的glnA mRNA。发现这两种mRNA的差异表达取决于氮源。glnB - A mRNA是固氮条件下的主要转录本，而glnA mRNA的合成几乎完全被抑制。glnA mRNA主要在含NH4(+)的培养基中产生。转录起始位点分析揭示了存在三种不同类型的氮调节启动子。GlnB - A mRNA从串联启动子选择性转录。其中一个类似于依赖NtrA的启动子，另一个类似于大肠杆菌的σ70启动子。glnA mRNA的合成由一个启动子调节，该启动子仅在固氮条件下（当分子氮是唯一氮源时）被抑制（或未被激活）。glnA前面的转录起始位点之前没有典型的大肠杆菌σ70启动子。在 - 33至 - 21位发现了一个类似于依赖NtrA的启动子共有序列的序列，只是存在一个基本错配。该序列与一个推定的“弱”NtrC结合位点重叠，类似于在肠道细菌中鉴定的那些位点。根据这些结果，推测glnA mRNA受一种新型氮调节启动子的控制。
