Suzuki I, Kikuchi H, Nakanishi S, Fujita Y, Sugiyama T, Omata T
Department of Applied Biological Sciences, School of Agricultural Sciences, Nagoya University, Japan.
J Bacteriol. 1995 Nov;177(21):6137-43. doi: 10.1128/jb.177.21.6137-6143.1995.
The gene (nirA) for nitrite reductase was cloned from the nonheterocystous, filamentous cyanobacterium Plectonema boryanum. The predicted protein consists of 654 amino acids and has a calculated molecular weight of 72,135. The deduced amino acid sequence from positions 1 to 511 is strongly similar to the entire sequence of the ferredoxin-dependent nitrite reductases from other phototrophs, while the remainder of the protein is unique to the Plectonema nitrite reductase. The C-terminal portion of the protein (amino acids 584 to 654) is 30 to 35% identical to [2Fe-2S] ferredoxins from higher plants and cyanobacteria, with all of the four Cys residues involved in binding of the [2Fe-2S] cluster in the ferredoxins being conserved. Immunoblotting analysis of the extracts of P. boryanum cells showed that the NirA polypeptide has an apparent molecular mass of 75 kDa. An insertional mutant of nirA lacked the 75-kDa polypeptide, had no nitrite reductase activity, and failed to grow on nitrate and nitrite, indicating that the novel nirA is the sole nitrite reductase gene in P. boryanum and that the NirA polypeptide with the ferredoxin-like domain is the apoprotein of the functional nitrite reductase. As in Synechococcus sp. strain PCC7942, nirA is the first gene of a large transcription unit (> 7 kb in size) and is repressed by ammonium and derepressed simply by deprivation of ammonium from the medium. The development of nitrite reductase activity was, however, found to require the presence of nitrate in the medium.
从非异形丝状蓝藻颤藻中克隆出了亚硝酸还原酶基因（nirA）。预测的蛋白质由654个氨基酸组成，计算分子量为72,135。从第1位到511位推导的氨基酸序列与其他光合生物中依赖铁氧化还原蛋白的亚硝酸还原酶的整个序列高度相似，而该蛋白质的其余部分是颤藻亚硝酸还原酶所特有的。该蛋白质的C末端部分（氨基酸584至654）与高等植物和蓝藻的[2Fe-2S]铁氧化还原蛋白有30%至35%的同一性，铁氧化还原蛋白中参与[2Fe-2S]簇结合的所有四个半胱氨酸残基都是保守的。对颤藻细胞提取物的免疫印迹分析表明，NirA多肽的表观分子量为75 kDa。nirA的插入突变体缺乏75 kDa的多肽，没有亚硝酸还原酶活性，并且不能在硝酸盐和亚硝酸盐上生长，这表明新的nirA是颤藻中唯一的亚硝酸还原酶基因，并且具有铁氧化还原蛋白样结构域的NirA多肽是功能性亚硝酸还原酶的脱辅基蛋白。与聚球藻属菌株PCC7942一样，nirA是一个大转录单元（大小>7 kb）的第一个基因，受铵抑制，仅通过从培养基中去除铵而解除抑制。然而，发现亚硝酸还原酶活性的发展需要培养基中存在硝酸盐。
