Andrews B J, Proteau G A, Beatty L G, Sadowski P D
Cell. 1985 Apr;40(4):795-803. doi: 10.1016/0092-8674(85)90339-3.
We have studied the interaction of purified FLP protein with restriction fragments from the substrate 2mu circle DNA of yeast. We find that FLP protects about 50 bp of DNA from nonspecific nuclease digestion. The protected site consists of two 13 bp inverted repeat sequences separated by an 8 bp spacer region. A third 13 bp element is also protected by binding of the FLP protein. We demonstrate that FLP introduces single- and double-strand breaks into the substrate DNA. This site-specific cleavage occurs at the margins of the spacer region, generating 8 bp 5' protruding ends with 5'-OH and 3'-protein-bound termini. Binding to mutant sites and half-sites demonstrates that the third symmetry element is not important for binding and cleavage by the FLP protein. The integrity of the core region is important for the cleavage activity of FLP.
我们研究了纯化的FLP蛋白与酵母底物2μm环状DNA的限制性片段之间的相互作用。我们发现FLP可保护约50bp的DNA不被非特异性核酸酶消化。受保护的位点由两个13bp的反向重复序列组成，中间间隔一个8bp的间隔区。第三个13bp元件也因FLP蛋白的结合而受到保护。我们证明FLP可在底物DNA中引入单链和双链断裂。这种位点特异性切割发生在间隔区的边缘，产生具有5'-OH和3'-蛋白结合末端的8bp 5'突出末端。与突变位点和半位点的结合表明，第三个对称元件对FLP蛋白的结合和切割并不重要。核心区域的完整性对FLP的切割活性很重要。
