Teebor G W, Frenkel K, Goldstein M S
Adv Enzyme Regul. 1982;20:39-54. doi: 10.1016/0065-2571(82)90007-3.
A methodology for the separation of radiation-induced thymine derivatives in DNA using high pressure liquid chromatography is presented. DNA was subjected to enzymatic hydrolysis yielding 2'-deoxyribonucleosides and the hydrolysate cochromatographed with marker compounds. Confirmation of the presence of derivatives was accomplished by chromatography on Sephadex LH-20 and microderivatization. The method separates free bases from nucleosides allowing for identification of spontaneously released bases or those released through the action of repair enzymes. The results indicate that most of the thymine derivatives formed in irradiated cellular DNA were the same as those found in DNA irradiated in solution. However, the major cellular derivative was not present in the latter. This derivative was identified as 5-hydroxymethyl-2'-deoxyuridine (HMdU). HMdU has previously been shown to be cytotoxic to cells in culture and caused diarrhea and bone marrow failure when administered to mice. Thus, the presence of this radiation-induced thymine derivative in cellular DNA correlates with the known effects of ionizing radiation on cells and animals.
本文介绍了一种使用高压液相色谱法分离DNA中辐射诱导胸腺嘧啶衍生物的方法。DNA经过酶促水解生成2'-脱氧核糖核苷，水解产物与标记化合物进行共色谱分析。通过在Sephadex LH-20上的色谱分析和微衍生化来确认衍生物的存在。该方法可将游离碱基与核苷分离，从而能够鉴定自发释放的碱基或通过修复酶作用释放的碱基。结果表明，辐照细胞DNA中形成的大多数胸腺嘧啶衍生物与溶液中辐照DNA中发现的衍生物相同。然而，后者中不存在主要的细胞衍生物。该衍生物被鉴定为5-羟甲基-2'-脱氧尿苷（HMdU）。先前已证明HMdU对培养细胞具有细胞毒性，给小鼠注射时会导致腹泻和骨髓衰竭。因此，细胞DNA中这种辐射诱导的胸腺嘧啶衍生物的存在与电离辐射对细胞和动物的已知影响相关。
