Kolb A, Busby S, Herbert M, Kotlarz D, Buc H
Institut Pasteur, Département de Biologie Moléculaire, Paris, France.
EMBO J. 1983;2(2):217-22. doi: 10.1002/j.1460-2075.1983.tb01408.x.
Polyacrylamide gel electrophoresis has been used to visualise and quantitate complexes between the Escherichia coli cyclic AMP receptor protein (CRP) and DNA fragments containing the promoter region of either the E. coli galactose or lactose operons. We show that, although CRP binding to the gal fragment is weaker than binding to the lac fragment, in each case, stable complexes are formed between one dimer of CRP and one molecule of DNA. We have examined the effects of a series of deletions and point mutations in the gal promoter region on CRP binding. From the position of deletions and mutations which prevent the formation of stable complexes, we deduce the location and extent of the sequence at the CRP binding site. We show that it covers approximately the same length of sequence as the binding site at the lac promoter. Unlike the lac site, the gal site contains no palindromic sequence. We discuss the importance of symmetry in the sequence at CRP binding sites and the validity of CRP binding consensus sequences which have been proposed.
聚丙烯酰胺凝胶电泳已被用于可视化和定量大肠杆菌环磷酸腺苷受体蛋白（CRP）与包含大肠杆菌半乳糖或乳糖操纵子启动子区域的DNA片段之间的复合物。我们表明，尽管CRP与gal片段的结合比与lac片段的结合弱，但在每种情况下，CRP的一个二聚体与一个DNA分子之间都会形成稳定的复合物。我们研究了gal启动子区域中一系列缺失和点突变对CRP结合的影响。从阻止形成稳定复合物的缺失和突变位置，我们推断出CRP结合位点处序列的位置和范围。我们表明，它覆盖的序列长度与lac启动子处的结合位点大致相同。与lac位点不同，gal位点不包含回文序列。我们讨论了CRP结合位点序列中对称性的重要性以及所提出的CRP结合共有序列的有效性。
