Simon T J, Smith C A, Friedberg E C
J Biol Chem. 1975 Nov 25;250(22):8748-52.
Previous studies have indicated that the ultraviolet endonuclease of bacteriophage T4 acts specifically at pyrimidine dimer sites in ultraviolet-irradiated DNA. At such sites the enzyme could conceivably catalyze endonucleolytic incision of the DNA either on the dimer-containing strand or on the strand directly opposite to the dimer. In the present work, a direct test of these alternatives was made. Substrate molecules containing one irradiated and one unirradiated strand were prepared from differentially isotopically labeled purified complementary strands of bacteriophage lambdaDNA. Following incubation with the enzyme, the sedimentation profiles of the DNA strands in alkaline sucrose density gradients were compared. The results show that the enzyme selectively nicks the irradiated strand.
先前的研究表明，噬菌体T4的紫外线内切核酸酶特异性作用于紫外线照射过的DNA中的嘧啶二聚体位点。在这些位点，该酶可以想象地催化DNA在含二聚体的链上或与二聚体直接相对的链上进行内切核酸酶切割。在本研究中，对这些可能性进行了直接测试。含有一条照射过的链和一条未照射过的链的底物分子是由噬菌体λDNA的差异同位素标记的纯化互补链制备的。在用该酶孵育后，比较了碱性蔗糖密度梯度中DNA链的沉降图谱。结果表明，该酶选择性地切割照射过的链。
