通过克隆的氧化亚铁硫杆菌ntrA基因对大肠杆菌σ54（NtrA）依赖性甲酸氢化酶活性的互补作用。
Complementation of Escherichia coli sigma 54 (NtrA)-dependent formate hydrogenlyase activity by a cloned Thiobacillus ferrooxidans ntrA gene.
作者信息
Berger D K, Woods D R, Rawlings D E
机构信息
Department of Microbiology, University of Cape Town, Rondebosch, South Africa.
出版信息
J Bacteriol. 1990 Aug;172(8):4399-406. doi: 10.1128/jb.172.8.4399-4406.1990.
The ntrA gene of Thiobacillus ferrooxidans was cloned by complementation of an Escherichia coli ntrA mutant that was unable to produce gas via the sigma 54 (NtrA)-dependent formate hydrogenlyase pathway. Analysis of the DNA sequence showed that the T. ferrooxidans ntrA gene coded for a protein of 475 amino acids (calculated Mr, 52,972). The T. ferrooxidans NtrA protein had 49, 44, 33, and 18% amino acid similarity with the NtrA proteins of Klebsiella pneumoniae, Azotobacter vinelandii, Rhizobium meliloti, and Rhodobacter capsulatus, respectively. The ability of the T. ferrooxidans NtrA protein to direct transcription from sigma 54-dependent promoters was demonstrated in E. coli by using fdhF-lacZ and nifH-lacZ fusions. An open reading frame coding for a protein of 241 amino acids (calculated Mr, 27,023) was situated 12 base pairs upstream of the T. ferrooxidans ntrA gene. Comparison of this protein with the product of the open reading frame ORF1, located upstream of the R. meliloti ntrA gene, showed that the two proteins had 55% amino acid similarity. The cloned T. ferrooxidans ntrA gene was expressed in E. coli from a promoter located within the ORF1 coding region.
通过对一株无法经由σ⁵⁴（NtrA）依赖的甲酸氢裂解酶途径产生气体的大肠杆菌ntrA突变体进行互补，克隆出了氧化亚铁硫杆菌的ntrA基因。DNA序列分析表明，氧化亚铁硫杆菌的ntrA基因编码一个由475个氨基酸组成的蛋白质（计算所得的分子量为52,972）。氧化亚铁硫杆菌的NtrA蛋白与肺炎克雷伯菌、维涅兰德固氮菌、苜蓿根瘤菌和荚膜红细菌的NtrA蛋白的氨基酸相似性分别为49%、44%、33%和18%。在大肠杆菌中，利用fdhF - lacZ和nifH - lacZ融合基因证明了氧化亚铁硫杆菌NtrA蛋白指导从σ⁵⁴依赖启动子进行转录的能力。一个编码由241个氨基酸组成的蛋白质（计算所得的分子量为27,023）的开放阅读框位于氧化亚铁硫杆菌ntrA基因上游12个碱基对处。将该蛋白与位于苜蓿根瘤菌ntrA基因上游的开放阅读框ORF1的产物进行比较，结果表明这两种蛋白的氨基酸相似性为55%。克隆的氧化亚铁硫杆菌ntrA基因在大肠杆菌中由位于ORF1编码区内的一个启动子进行表达。
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