Morgan T V, Lundell D J, Burgess B K
Corporate Research Laboratory, EXXON Research and Engineering Company, Annandale, New Jersey 08801.
J Biol Chem. 1988 Jan 25;263(3):1370-5.
The structure of Azotobacter vinelandii ferredoxin I (AvFdI) has been extensively characterized by a variety of techniques. Although its physiological function is unknown, it has long been implicated as being involved in electron donation to nitrogenase. Here we report that the AvFdI gene (fdxA) has been cloned from an EcoRI digest lambda library using a synthetic oligonucleotide probe and that its sequence has been determined. The amino acid sequence deduced from the DNA sequence is identical to the previously published protein sequence. Analysis of the promoter region indicates that AvFdI is not a nif specific gene product. A mutant of A. vinelandii has been constructed which is identical to the wild-type, at the DNA level, except that the fdxA gene has been interrupted by insertion of a kanamycin cartridge. This mutant, called LM100, does not synthesize AvFdI but does synthesize the Fe and MoFe proteins of nitrogenase and grows at wild-type rates under N2-fixing conditions. This demonstrates that AvFdI is not required for N2 fixation by A. vinelandii. There is a small acidic protein, which is present in wild-type A. vinelandii, whose level is dramatically increased in LM100. The nature of this protein is under further investigation.
维涅兰德固氮菌铁氧化还原蛋白I（AvFdI）的结构已通过多种技术进行了广泛表征。尽管其生理功能尚不清楚，但长期以来一直认为它参与向固氮酶的电子供体过程。在此我们报告，已使用合成寡核苷酸探针从EcoRI消化的λ文库中克隆出AvFdI基因（fdxA），并确定了其序列。从DNA序列推导的氨基酸序列与先前发表的蛋白质序列相同。对启动子区域的分析表明，AvFdI不是一个nif特异性基因产物。构建了一种维涅兰德固氮菌突变体，在DNA水平上它与野生型相同，只是fdxA基因因插入卡那霉素盒而被中断。这个名为LM100的突变体不合成AvFdI，但能合成固氮酶的Fe和MoFe蛋白，并在固氮条件下以野生型速率生长。这表明维涅兰德固氮菌进行固氮作用不需要AvFdI。在野生型维涅兰德固氮菌中存在一种小的酸性蛋白，其水平在LM100中显著增加。这种蛋白的性质正在进一步研究中。
