Otani M, Hwang H S, Nakazawa T, Wataya Y
Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Nucleic Acids Symp Ser. 1992(27):193-4.
The mechanism of intracellular deoxyribonucleotide triphosphates (dNTP) pool imbalance-induced cell death in mouse FM3A cells was studied. When the cells were treated with 1 microM 5-fluorodeoxyuridine (FdUrd), the imbalance of the cellular dNTP pool was induced. The imbalance was followed by DNA double stranded breaks and subsequent cell death. The endonuclease toward double stranded DNA has been found in a fraction of FdUrd treated cell lysate, and isolated using column chromatography. SDS-polyacrylamide gel electrophoresis showed a major protein species of approximate 45 kDa. The endonuclease was revealed, using electrophoretic separation in SDS-polyacrylamide gels containing DNA, by incubating the gels in buffer to remove SDS and to allow renaturation and enzyme activity.
研究了小鼠FM3A细胞中细胞内脱氧核糖核苷酸三磷酸（dNTP）池失衡诱导细胞死亡的机制。当用1微摩尔5-氟脱氧尿苷（FdUrd）处理细胞时，会诱导细胞dNTP池失衡。随后出现DNA双链断裂及细胞死亡。在一部分经FdUrd处理的细胞裂解物中发现了针对双链DNA的核酸内切酶，并通过柱色谱法进行分离。SDS-聚丙烯酰胺凝胶电泳显示主要蛋白质条带约为45 kDa。通过在含有DNA的SDS-聚丙烯酰胺凝胶中进行电泳分离，将凝胶在缓冲液中孵育以去除SDS并使其复性和恢复酶活性，从而揭示该核酸内切酶。
