Galinier A, Garnerone A M, Reyrat J M, Kahn D, Batut J, Boistard P
Laboratoire de Biologie Moléculaire des Relations Plantes-Microorganismes, INRA-CNRS, Castanet-Tolosan, France.
J Biol Chem. 1994 Sep 23;269(38):23784-9.
The FixJ protein is a member of the regulator class of two-component systems involved in the transcriptional activation of nitrogen fixation genes in Rhizobium meliloti. Phosphorylation of FixJ was previously demonstrated to dramatically enhance its transcriptional activity at the nifA and fixK promoters. Here we show that the isolated carboxyl-terminal domain of FixJ, FixJC, binds the fixK promoter, whereas binding of the full-length FixJ protein requires its phosphorylation. By analyzing the DNase I and Exonuclease III protection patterns of the wild-type and a mutant fixK promoter, we have identified two overlapping binding regions for both phosphorylated FixJ and FixJC. A higher affinity region is located between positions -69 and -44 relative to the transcription start site, and a lower affinity region, between positions -57 and -31, overlaps the -35 region of the promoter.
FixJ蛋白是参与苜蓿根瘤菌固氮基因转录激活的双组分系统调节类成员。先前已证明FixJ的磷酸化可显著增强其在nifA和fixK启动子处的转录活性。在此我们表明，分离出的FixJ羧基末端结构域FixJC可结合fixK启动子，而全长FixJ蛋白的结合则需要其磷酸化。通过分析野生型和突变型fixK启动子的DNase I和核酸外切酶III保护模式，我们确定了磷酸化FixJ和FixJC的两个重叠结合区域。相对于转录起始位点，较高亲和力区域位于-69至-44位之间，较低亲和力区域位于-57至-31位之间，与启动子的-35区域重叠。
