一种识别DNA中环丁烷型胸腺嘧啶二聚体的单克隆抗体的建立：与针对（6-4）光产物的64M-1抗体的比较研究

Establishment of a monoclonal antibody recognizing cyclobutane-type thymine dimers in DNA: a comparative study with 64M-1 antibody specific for (6-4)photoproducts.

作者信息

Mizuno T, Matsunaga T, Ihara M, Nikaido O

机构信息

Division of Radiation Biology, Faculty of Pharmaceutical Sciences, Kanazawa University, Japan.

出版信息

Mutat Res. 1991 Mar;254(2):175-84. doi: 10.1016/0921-8777(91)90009-e.

DOI:10.1016/0921-8777(91)90009-e

PMID:2002812

Abstract

We obtained a monoclonal antibody (TDM-1) binding to 313-nm UV-irradiated DNA in the presence of acetophenone. The binding of TDM-1 to 254-nm UV-irradiated DNA was not reduced with the subsequent irradiation of 313-nm UV. Furthermore, the treatment of UV-irradiated DNA with photolyase from E. coli and visible light exposure reduced both the antibody binding and the amount of thymine dimers in the DNA. A competitive inhibition assay revealed that the binding of TDM-1 to UV-irradiated DNA was inhibited with photolyase, but not with 64M-1 antibody specific for (6-4)photoproducts. These results suggest that TDM-1 antibody recognizes cyclobutane-type thymine dimers in DNA. Using TDM-1 and 64M-1 antibodies, we differentially measured each type of damage in DNA extracted from UV-irradiated mammalian cells. Repair experiments confirm that thymine dimers are excised from UV-irradiated cellular DNA more slowly than (6-4)photoproducts, and that the excision rates of thymine dimers and (6-4)photoproducts are lower in mouse NIH3T3 cells than in human cells.

摘要

我们获得了一种单克隆抗体（TDM-1），它在苯乙酮存在的情况下与313纳米紫外线照射的DNA结合。TDM-1与254纳米紫外线照射的DNA的结合在随后用313纳米紫外线照射时并未减少。此外，用来自大肠杆菌的光裂解酶处理紫外线照射的DNA并暴露于可见光下，可减少抗体结合以及DNA中胸腺嘧啶二聚体的数量。竞争性抑制试验表明，TDM-1与紫外线照射的DNA的结合被光裂解酶抑制，但不被对（6-4）光产物特异的64M-1抗体抑制。这些结果表明，TDM-1抗体识别DNA中的环丁烷型胸腺嘧啶二聚体。使用TDM-1和64M-1抗体，我们分别测量了从紫外线照射的哺乳动物细胞中提取的DNA中的每种损伤类型。修复实验证实，胸腺嘧啶二聚体从紫外线照射的细胞DNA中切除的速度比（6-4）光产物慢，并且在小鼠NIH3T3细胞中胸腺嘧啶二聚体和（6-4）光产物的切除率低于人类细胞。