Plumbridge J, Kolb A
Institut de Biologie Physico-chimique (UPR9073), 13 rue P. et M. Curie, 75005 Paris, France.
Nucleic Acids Res. 1998 Mar 1;26(5):1254-60. doi: 10.1093/nar/26.5.1254.
Repression of the divergent nagE - B operons requires NagC binding to two operators which overlap the nagE and nagB promoters, resulting in formation of a DNA loop. Binding of the cAMP/CAP activator to its site, adjacent to the nagE operator, stabilizes the DNA loop in vitro. The DNA of the nagE-B intergenic region is intrinsically bent, with the bend centred on the CAP site. We show that displacement of the CAP site by 6 bp results in complete derepression of the two operons. This derepression is observed even in the absence of cAMP/CAP binding and despite the fact that the two NagC operators are still in phase, demonstrating that the inherently bent structure of the DNA loop is important for repression. Since no interaction between NagC and CAP has been detected, we propose that the role of CAP in the repression loop is architectural, stabilizing the intrinsic bend. The cAMP/CAP complex is necessary for activation of the nagE-B promoters. In this case protein-protein contacts between CAP and RNA polymerase are necessary for full activation, but at least a part of the activation is likely due to an effect of CAP binding altering DNA structure.
对反向排列的nagE - B操纵子的阻遏需要NagC结合到两个与nagE和nagB启动子重叠的操纵基因上，从而形成一个DNA环。cAMP/CAP激活剂与其位点（邻近nagE操纵基因）的结合在体外稳定了DNA环。nagE - B基因间区域的DNA具有内在弯曲，弯曲中心位于CAP位点。我们发现，将CAP位点移位6个碱基对会导致这两个操纵子完全去阻遏。即使在没有cAMP/CAP结合的情况下，并且尽管两个NagC操纵基因仍保持相位，这种去阻遏现象依然会出现，这表明DNA环的固有弯曲结构对阻遏很重要。由于未检测到NagC与CAP之间的相互作用，我们提出CAP在阻遏环中的作用是构建性的，即稳定固有弯曲。cAMP/CAP复合物对于nagE - B启动子的激活是必需的。在这种情况下，CAP与RNA聚合酶之间的蛋白质 - 蛋白质接触对于完全激活是必需的，但至少部分激活可能是由于CAP结合改变DNA结构的作用。
