Arsene F, Kaminski P A, Elmerich C
Centre National de la Recherche Scientifique Unité Recherche Associée 1300. Département des Biotechnologies, Institut Pasteur, Paris, France.
J Bacteriol. 1996 Aug;178(16):4830-8. doi: 10.1128/jb.178.16.4830-4838.1996.
Azospirillum brasilense NifA, which is synthesized under all physiological conditions, exists in an active or inactive from depending on the availability of ammonia. The activity also depends on the presence of PII, as NifA is inactive in a glnB mutant. To investigate further the mechanism that regulates NifA activity, several deletions of the nifA coding sequence covering the amino-terminal domain of NifA were constructed. The ability of these truncated NifA proteins to activate the nifH promoter in the absence or presence of ammonia was assayed in A. brasilense wild-type and mutant strains. Our results suggest that the N-terminal domain is not essential for NifA activity. This domain plays an inhibitory role which prevents NifA activity in the presence of ammonia. The truncated proteins were also able to restore nif gene expression to a glnB mutant, suggesting that PII is required to activate NifA by preventing the inhibitory effect of its N-terminal domain under conditions of nitrogen fixation. Low levels of nitrogenase activity in the presence of ammonia were also observed when the truncated gene was introduced into a strain devoid of the ADP-ribosylation control of nitrogenase. We propose a model for the regulation of NifA activity in A. brasilense.
巴西固氮螺菌NifA在所有生理条件下均有合成，其以活性或非活性形式存在，这取决于氨的可利用性。其活性还取决于PII的存在，因为NifA在glnB突变体中无活性。为了进一步研究调节NifA活性的机制，构建了多个覆盖NifA氨基末端结构域的nifA编码序列缺失体。在巴西固氮螺菌野生型和突变株中检测了这些截短的NifA蛋白在有无氨存在的情况下激活nifH启动子的能力。我们的结果表明，N末端结构域对NifA活性不是必需的。该结构域起抑制作用，在有氨存在时阻止NifA活性。截短的蛋白还能够恢复glnB突变体中的nif基因表达，这表明在固氮条件下，PII通过阻止其N末端结构域的抑制作用来激活NifA。当将截短基因导入缺乏固氮酶ADP-核糖基化调控的菌株中时，在有氨存在的情况下也观察到了低水平的固氮酶活性。我们提出了一个巴西固氮螺菌中NifA活性调节的模型。
