Gubler M, Hennecke H
Mikrobiologisches Institut, ETH-Zentrum, Zürich, Switzerland.
J Bacteriol. 1988 Mar;170(3):1205-14. doi: 10.1128/jb.170.3.1205-1214.1988.
The transcriptional start site of the Bradyrhizobium japonicum fixBC operon was identified by nuclease S1 mapping. It was located approximately 700 base pairs upstream of fixB and was preceded by a promoter sequence that showed strong homology to the B. japonicum fixA promoter and thus to the general nif consensus promoter sequence. Further transcript mapping experiments revealed that fixA and fixBC transcription in B. japonicum strictly depended on the presence of the regulatory gene nifA and on low oxygen partial pressure. Consistent with these data, chromosomally integrated fixA- and fixB-lacZ fusions expressed beta-galactosidase activity only in the wild type but not in a nifA mutant and only under microaerobic but not aerobic growth conditions. The presence of nifA accounted for a 19-fold and 44-fold activation of the fixA and fixB promoters, respectively. These results show that the fixA and fixBC genes are regulated in a way similar to that of the nitrogenase genes nifH and nifDK. A very peculiar finding was that the fixA and fixB promoters, when they were located on plasmids, could hardly be activated by the NifA protein, irrespective of whether this was tested in Escherichia coli or B. japonicum backgrounds. This is in clear contrast to the situation with nifH and nifD promoters.
通过核酸酶S1图谱分析确定了慢生根瘤菌固氮基因fixBC操纵子的转录起始位点。它位于fixB上游约700个碱基对处，其前面的启动子序列与慢生根瘤菌fixA启动子具有高度同源性，因此与一般的固氮基因一致序列启动子也具有高度同源性。进一步的转录图谱实验表明，慢生根瘤菌中fixA和fixBC的转录严格依赖于调控基因nifA的存在以及低氧分压。与这些数据一致的是，染色体整合的fixA-和fixB-lacZ融合蛋白仅在野生型中表达β-半乳糖苷酶活性，而在nifA突变体中不表达，并且仅在微需氧而非好氧生长条件下表达。nifA的存在分别使fixA和fixB启动子的活性提高了19倍和44倍。这些结果表明，fixA和fixBC基因的调控方式与固氮酶基因nifH和nifDK相似。一个非常奇特的发现是，当fixA和fixB启动子位于质粒上时，无论在大肠杆菌还是慢生根瘤菌背景中进行测试，它们几乎都不能被NifA蛋白激活。这与nifH和nifD启动子的情况形成了鲜明对比。
