Yields of damage to C4' deoxyribose and to pyrimidines in pUC18 by the direct effect of ionizing radiation.
Yields of damage to C4' deoxyribose and to pyrimidines in pUC18 by the direct effect of ionizing radiation.
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电离辐射的直接作用对 pUC18 中的 C4 脱氧核糖和嘧啶造成损害。
DOI：
10.1093/nar/gks271
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发表时间：
2012
影响因子：
14.9
通讯作者：
Bernhard,WilliamA
中科院分区：
文献类型：
--
作者：
Peoples,AnitaR;Lee,Jane;Weinfeld,Michael;Milligan,JamieR;Bernhard,WilliamA
我们对DNA损伤形成的直接作用的机械理解在很大程度上依赖于EPR光谱研究的自由基中间体。将该信息与稳定产物形成相联系需要具有可比灵敏度的方法，这是由Weinfeld和Soderlind开发的32 P-后标记测定法所满足的标准[Weinfeld，M. Soderlind，K. J.M.（1991）32 P-Postlabeling detection of radiation-induced DNA damage：identification and estimation of thymine glycol and phosphoglycolate termini.Biochemistry，30，1091-1097]，当应用于间接效应时，检测到了磷酸乙醇酸（pg）和胸腺嘧啶二醇（Tg）。在此，我们将该测定应用于直接效应，测量具有2.5、16或23沃茨/核苷酸的水合水平（Γ）的PUC 18膜中的产物产率，并在4K或室温（RT）下进行X辐照。在RT和4K条件下，Γ_2.5的pg [G（pg）]产率分别为2.8 ± 0.2nmol/J和0.2 ± 0.3nmol/J，表明C_4 ′自由基对总脱氧核糖损伤的直接作用很小。在Γ = 2.5时，可检测到的碱基损伤产率[G（B*）]为30.2 ± 1.0 nmol/J（RT）和12.9 ± 0.7 nmol/J（4 K）。虽然碱基损伤称为B*，可能是由于氧化或还原，我们认为，两个还原产物，5，6-二氢尿嘧啶和5，6-二氢胸腺嘧啶，是最有可能的候选人。
Our mechanistic understanding of damage formation in DNA by the direct effect relies heavily on what is known of free radical intermediates studied by EPR spectroscopy. Bridging this information to stable product formation requires methods with comparable sensitivities, a criterion met by the32P-post-labeling assay developed by Weinfeld and Soderlind, [Weinfeld,M. and Soderlind,K.-J.M. (1991)32P-Postlabeling detection of radiation-induced DNA damage: identification and estimation of thymine glycols and phosphoglycolate termini.Biochemistry, 30, 1091–1097] which when applied to the indirect effect, detected phosphoglycolate (pg) and thymine glycol (Tg). Here we applied this assay to the direct effect, measuring product yields in pUC18 films with hydration levels (Γ) of 2.5, 16 or 23 waters per nucleotide and X-irradiated at either 4 K or room temperature (RT). The yields of pg [G(pg)] for Γ ∼ 2.5 were 2.8 ± 0.2 nmol/J (RT) and 0.2 ± 0.3 nmol/J (4 K), which is evidence that the C4′ radical contributes little to the total deoxyribose damage via the direct effect. The yield of detectable base damage [G(B*)] at Γ ∼ 2.5 was found to be 30.2 ± 1.0 nmol/J (RT) and 12.9 ± 0.7 nmol/J (4 K). While the base damage called B*, could be due to either oxidation or reduction, we argue that two reduction products, 5,6-dihydrouracil and 5,6-dihydrothymine, are the most likely candidates.