Hill S A, Samuels D S, Carlson J H, Wilson J, Hogan D, Lubke L, Belland R J
Laboratory of Microbial Structure and Function, National Institutes of Health, National Institutes of Allergy and Infectious Diseases, Rocky Mountain Laboratories, Hamilton, Montana 59840, USA.
Mol Microbiol. 1997 Feb;23(4):649-56. doi: 10.1046/j.1365-2958.1997.2321612.x.
Integration host factor (IHF) is a small, heterodimeric DNA-binding protein with pleiotropic function. IHF was purified to apparent homogeneity from Neisseria gonorrhoeae. Gel-retardation assays demonstrated binding of IHF to the pilE promoter region. The IHF-binding site was identified by DNase I protection assays and mapped proximal to three previously defined pilE promoters. Removal of the putative IHF-binding domain from pilE promoter DNA negated retardation of the DNA fragment when assessed by gel-shift analysis. Kleinschmidt electron microscopy showed pronounced kinking of pilE promoter DNA following incubation with IHF. Isogenic N. gonorrhoeae strains were constructed that contained either a wild-type pilE locus or a deleted pilE locus where the IHF-binding domain was removed. Primer-extension analysis and Northern blotting of total gonococcal RNA showed that in the absence of IHF binding at the pilE promoter, transcription was reduced 10-fold. Together, these data indicate that IHF is a transcriptional co-activator of pilE.
整合宿主因子(IHF)是一种具有多效功能的小型异源二聚体DNA结合蛋白。IHF从淋病奈瑟菌中纯化至表观均一。凝胶阻滞分析表明IHF与pilE启动子区域结合。通过DNA酶I保护分析鉴定了IHF结合位点,并将其定位在三个先前定义的pilE启动子附近。通过凝胶迁移分析评估时,从pilE启动子DNA中去除假定的IHF结合结构域可消除DNA片段的阻滞。克莱因施密特电子显微镜显示,与IHF孵育后,pilE启动子DNA出现明显的扭结。构建了同基因的淋病奈瑟菌菌株,其包含野生型pilE基因座或缺失了IHF结合结构域的pilE基因座。对总淋球菌RNA进行引物延伸分析和Northern印迹分析表明,在pilE启动子处不存在IHF结合时,转录减少了10倍。这些数据共同表明,IHF是pilE的转录共激活因子。