Dna Radiation Damage and its Modification by Metallothionein

Thiol compounds have long been known to protect living cells against the harmful effects of ionizing radiation. Maetallothionein is a naturally occurring low molecular weight polypeptide rich in cysteine residues and may be useful in protection against low-level radiation effects.

Radiation damage to DNA and its nucleotide components and the radioprotective effect of metallothionein have been studied in model chemical systems and compared to its effect on cells. Metallothionein acts both as a free radical scavenger and a reductant, and its radioprotective effectiveness has been studied as a function of dose, drug concentration, and in the presence and absence of oxygen. It is more effective in protecting against sugar-phosphate damage under hypoxic conditions. The chemical modification is greater than that of cell killing as measured by the loss of colony-forming ability. Dose reduction factors greater than two are observed for DNA radioprotection, but the values in cells are much lower. These findings will be discussed in terms of the molecular mechanisms and their implications.     

Dna Damage by Chemically Generated Singlet Oxygen

A naphthalenic endoperoxide was used as a non-photochemical source of singlet oxygen (¹O₂) to examine some interactions between this reactive oxygen species and DNA. High molecular weight DNA (ca. 10⁸ daltons) was exposed to 120 mol m^?31O₂ (cumulative concentration) and analyzed for interstrand crosslinkage by hydroxyl apatite chromatography following formamide denaturation. No evidence for ¹O₂-induced interstrand crosslinking was obtained. The capacity of ¹O₂ to generate strand breaks in single-stranded (ss) and double-stranded (ds) DNA was investigated by sucrose gradient centrifugation analysis of bacteriophage øX174 DNA. No direct strand breaks could be detected at neutral pH, whereas extensive strand breakage was observed after treatment with alkali. Possible biological consequences of ¹O₂ -exposure were assessed by examining the plaque-forming capacity of ss and ds øX 174 DNA molecules using wildtype Escherichia coli spheroplasts as recipients. Without any further treatment with heat or alkali, exposure to the endoperoxide resulted in a time- and dose-dependent inactivation, ss DNA being considerably more sensitive than ds DNA. From the present results and those reported earlier (Nieuwint et al.,²⁰) we infer that ¹O₂-induced inactivation of øX174 DNA is not due to DNA backbone breakage nor to interstrand crosslinking, but rather to some form of damage to the base or sugar moiety of the DNA, the exact nature of which remains to be elucidated.     

SUMO化修饰在DNA损伤响应中的作用

物理或化学等多种因素均可以引起DNA损伤。为维持机体基因组的稳定性,机体形成了精确完整的机制来修复损伤的／DNA。SUMO（smallubiquitin-relatedmodifier,SUMO）化修饰与其他蛋白翻译后修饰一样,具有多种生物学功能。近年来的研究表明,其在DNA损伤修复中也具有非常重要的作用。该文就DNA损伤修复、SUMO,96修饰系统及其二者关系的最新研究进展作了较为全面的介绍和总结。     

蓝藻的类金属硫蛋白及其研究进展

单细胞蓝藻中的类金属硫蛋白已经得到分离纯化,并在蛋白质水平上与标准的哺乳动物金属硫蛋白做了对比性分析,发现二者氨基酸组成和序列差异很大,前者只形成一个结构域,但二级结构和金属结合性质具有一定的相似性,是进化上功能趋同的表现．同时克隆并分析了它的基因ORF结构,研究了金属诱导和逆向转录抑制对于蛋白质表达的调控及类似于哺乳动物金属硫蛋白基因具有的放大和重排现象机理,提出了近期研究的重点和方向．     

32P-Postlabeling Assay for Free Radical-Induced Dna Damage: The Formamido Remnant of Thymine

A prominent lesion in DNA exposed to oxidative free radicals results from the degradation of thymine leaving a formamido remnant. A ³²P-postlabeling assay has been developed for the detection of the formamido lesion. The assay is based on the circumstance that the lesion prevents hydrolysis by nuclease PI of the phosphoester bond 3' to the damaged nucleoside. Thus, a nuclease PI plus acid phosphatase digest of DNA generates mostly nucleosides whereas the formamido lesion is rendered as a modified dinucleoside monophosphate. Dinucleoside monophosphates, but not nucleosides, are apt substrates for ³²P-postlabeling by polynucleotide kinase. The assay was applied to calf thymus DNA X-irradiated in oxygenated solution. The formamido lesion could be detected down to a dose of a few Gy.     

Bleomycin-Iron Damage To Dna With Formation Of 8-Hydroxydeoxyguanosine and Base Propenals. Indications That Xanthine Oxidase Generates Superoxide From Dna Degradation Products

Bleomycin, in the presence of ferric salts, oxygen and a suitable reductant, degrades DNA with the release of base propenals, detected as thiobarbituric acid (TBA) reactivity, and the formation of 8-hydroxydeo-xyguanosine (80HdG) detected by HPLC. When xanthine oxidase is added to the incubated mixture of DNA degradation products, TBA-reactivity is destroyed but 80HdG formation is increased. EPR Spin trapping experiments show that hydroxyl radicals (OH) are formed in the reaction mixture and can be inhibited by the inclusion of either superoxide dismutase or catalase. These findings suggest that the base propenals and possibly malondialdehyde, formed from them, are aldehydic substrates for xanthine oxidase and, the product of this reaction is superoxide (O₂^-) and hydrogen peroxide (H₂O₂). Thus, TBA reactivity is destroyed in the formation of O₂^- and H₂O₂ which stimulate further oxidative damage to DNA resulting in increased 8OHdG formation.     

紫外辐射诱导烟草原生质体中DNA损伤的彗星电泳检测

以烟草原生质体为材料,采用彗星电泳检测用0.5W·m^-2紫外线以不同时间（0、5、10、30、60和120s）诱导的烟草原生质体中DNA的损伤。结果表明,在0～10s的时间内代表DNA损伤程度的尾矩、Olive尾矩等参数与紫外线照射时间具有良好的时间依赖关系。本文建立的烟草原生质体体系采用彗星电泳技术,可以快速而灵敏地检测紫外线对植物细胞的损伤程度。     

单克隆抗体ELISA测定UVB诱导的DNA损伤

本文采用单克隆抗体酶联免疫吸附分析法测定了ＵＶ－Ｂ诱导ＤＮＡ产生的ＣＰＤ和６－４ＰＰ。经０．５ｍＷ／ｃｍ＾２ＵＶ－Ｂ处理１５ｍｉｎ的小牛胸物鲱鱼精ＤＮＡ,ＣＰＤ和６－４ＰＰ含量显著增加,而未经ＵＶ－Ｂ处理的对照ＤＮＡ则没有二聚体形成。     

单克隆抗体ELISA测定UVB诱导的DNA损伤(英文)

本文采用单克隆抗体酶联免疫吸附分析法测定了ＵＶＢ诱导ＤＮＡ产生的ＣＰＤ和６４ＰＰ。经０．５ｍＷ／ｃｍ２ＵＶＢ处理１５ｍｉｎ的小牛胸腺和鲱鱼精ＤＮＡ,ＣＰＤ和６４ＰＰ含量显著增加,而未经ＵＶＢ处理的对照ＤＮＡ则没有二聚体形成。     

UV-B辐射和蒽对三角褐指藻DNA伤害的相互作用

运用生态毒理学和生物化学的方法研究了紫外线和多环芳烃－蒽对三角褐指藻DNA的伤害作用,结果表明,蒽对三角褐指藻的生长有抑制作用;随着蒽浓度的增加,三角揭褐藻DNA损伤程度增加;在蒽浓度固定不变时,随着处理时间的延长,DNA的损伤程度同样提高;在蒽的处理过程中同时伴有紫外线的辐射处理,DNA的损伤程度加剧;蒽处理解除一段时间后,DNA损伤程度未明显减轻、而UV－B处理解除后,DNA的损伤可明显恢复,说明DNA的损伤可在一定程度上指示海洋微藻受蒽伤的程度。     

DNA Damage Checkpoint, Damage Repair, and Genome Stability

Genomic DNA is under constant attack from both endogenous and exogenous sources of DNA damaging agents. Without proper care, the ensuing DNA damages would lead to alteration of genomic structure thus affecting the faithful transmission of genetic information. During the process of evolution, organisms have acquired a series of mechanisms responding to and repairing DNA damage, thus assuring the maintenance of genome stability and faithful transmission of genetic information. DNA damage checkpoint is one such important mechanism by which, in the face of DNA damage, a cell can respond to amplified damage signals, either by actively halting the cell cycle until it ensures that critical processes such as DNA replication or mitosis are complete or by initiating apoptosis as a last resort. Over the last decade, complex hierarchical interactions between the key components like ATM/ATR in the checkpoint pathway and various other mediators, effectors including DNA damage repair proteins have begun to emerge. In the meantime, an intimate relationship between mechanisms of damage checkpoint pathway, DNA damage repair, and genome stability was also uncovered. Reviewed hereinare the recent findings on both the mechanisms of activation of checkpoint pathways and their coordination with DNA damage repair machinery as well as their effect on genomic integrity.     

DNA损伤检验点与损伤修复及基因组稳定性

生物有机体基因组DNA经常会受到内源或外源因素的影响而导致结构发生变化,产生损伤;在长期进化过程中,有机体也相应形成了一系列应对与修复损伤DNA,并维持染色体基因组正常结构功能的机制。其中DNA损伤检验点（DNA damage checkpoint）就是在感应DNA损伤的基础上,对损伤感应信号进行转导,或引起细胞周期的暂停,从而使细胞有足够的时间对损伤DNA进行修复,或最终导致细胞发生凋亡。DNA损伤检验点信号转导途径是一个高度保守的信号感应过程,整个途径大致可以分为损伤感应、信号传递及信号效应3个组成部分。其中3-磷脂酰肌醇激酶家族类成员ATM（ataxia-telangiectasia mutated）和ATR（ataxia-telangiectasia and Rad3-related）活性的增加构成整个途径活化的第一步。它们通过激活下游的效应激酶,Chk2／Chk1,通过协同作用许多其他调控细胞周期、DNA复制、DNA损伤修复及细胞凋亡等过程的蛋白质因子来实现细胞对DNA损伤的高度协调反应。近十几年,随着此领域研究的不断深入,人们逐步揭示了DNA损伤检验点途径发生过程中,各种核心组分通过与不同调节因子、效应因子及DNA损伤修复蛋白间的复杂相互作用,以实现监测感应异常DNA结构并实施相应反应的机制;其中,检验点衔接因子（mediators）及染色质结构,尤其是核小体组蛋白的共价修饰在调控ATM／ATR活性,促进ATM／ATR与底物间的相互作用以及介导DNA损伤位点周围染色质区域上多蛋白复合物在时间与空间上的动态形成发挥着重要的作用。同时,人们也开始发现DNA损伤检验点途径与DNA损伤修复、基因组稳定性以及肿瘤发生等过程之间某些内在的联系。该反应途径在通过协调细胞针对DNA损伤做出各种反应的基础上,直接或间接地参与或调控DNA损伤修复过程,并与DNA损伤修复途径协同作用最终保证染色体基凶组结构的完整性,而检验点途径的改变,则会引起基因组不稳定的发生,包括从突变频率的提高到大范围的染色体重排,以及染色体数量的畸变。如：突变发生在肿瘤形成早期,会大大增加肿瘤发生的几率。文章将对DNA损伤检验点途径机制及其对DNA损伤修复、基因组稳定性影响的最新进展进行综述。     

Enhancement of Nitrogen Dioxide-Induced Lipid Peroxidation and Dna Strand Breaking by Cysteine and Glutathione

Nitrogen dioxide less than 100 ppm in air induced lipid peroxidation of liposome composed of l-palmitoyl-2-arachidonylphosphatidylcholine as assessed by thiobarbituric acid reactivity. The nitrogen dioxide-induced lipid peroxidation was enhanced by cysteine, glutathione and bovine serum albumin. While the activity of nitrogen dioxide in air to induce single strand breaks of supercoiled plasmid DNA was low, the breaking was remarkably enhanced by cysteine, glutathione and bovine serum albumin. ESR spin trapping using 5,5-dimethyl-1-pyrroline N-oxide showed that certain strong oxidant(s) were generated by interaction of nitrogen dioxide and cysteine. The spin trapping using 3,5-dibromo-4-nitrosobenzene-sulfonate suggested that sulfur-containing radicals were generated by interaction of nitrogen dioxide and cysteine or glutathione. Hence, certain sulfur-containing radicals generated by the interaction which could effectively induce lipid peroxidation and DNA strand breaks.     

病毒感染与宿主细胞DNA损伤应答

病毒感染宿主细胞后,利用细胞内的营养物质和原料进行复制和增殖,同时能引起宿主细胞启动抗病毒免疫应答的防御机制。此外,近年来的研究还表明病毒感染能够引起宿主细胞的DNA损伤应答,该反应是细胞另一种防止病毒入侵的自我保护机制。同时发现,病毒在长期进化过程中形成了不同的机制来对抗宿主细胞的DNA损伤应答,从而消除细胞对其复制和繁殖产生的不利影响。因此,研究和阐述病毒感染后引起宿主细胞DNA损伤应答途径的机制,可使我们采取相应对策选择新的抗病毒靶点,从而有利于新型抗病毒药物的开发。     

自由基对线粒体DNA的氧化损伤与衰老

自由基是一类氧化剂,对生物具有多种损害作用．衰老的自由基学说是有关衰老机理的诸多学说之一．线粒体DNA组成结构特殊,易受自由基攻击;目前认为,线粒体DNA的氧化损伤是自由基引起衰老的分子基础．     

调控DNA损伤修复基因的微小RNA

随着对DNA损伤修复基因研究的深入,其信号转导路径及调控网络也进一步明了,调控DNA损伤修复基因的微小RNA(miRNA)也越来越多地被认识和发现。简要综述了DNA损伤途径中调控主要的损伤修复基因的miRNA,有助于深入阐明DNA损伤修复机制,为开发抗辐射药物和临床上DNA损伤修复异常相关肿瘤的基因治疗提供新的靶点。     

盐穗木金属硫蛋白HcMT的体外自由基清除活性及抗氧化能力*

目的: 原核表达盐穗木(Halostachys caspica C. A. Mey.)金属硫蛋白HcMT并探究其抗氧化活性。方法: 构建原核表达载体pET-32a-HcMT,转化至大肠杆菌Escherichia coli BL21,加入Zn²⁺胁迫培养(终浓度为200 μmol/L),分离纯化得到Zn-HcMT,测定Zn-HcMT自由基清除活性和总抗氧化能力,制备复合物Zn-HcMT/TiO₂并做FTIR表征。结果: 通过原核表达获得融合蛋白Zn-HcMT,对·OH、O₂·^-、DPPH自由基具有较强的清除活性,对·OH、O₂·^-的IC₅₀分别为0.386 mg/mL、0.038 mg/mL。融合蛋白浓度为0.01 mg/mL时,对DPPH清除率达(37.43 ± 0.006 8)%,浓度为0.3mg/mL时TEAC(trolox-equivalent antioxidant capacity)值为(1.023 ± 0.01)mmol/L,融合蛋白还原力A₇₀₀为0.142 ± 0.055,FTIR图谱同时表现了Zn-HcMT和TiO₂吸收特性。结论: Zn-HcMT具有良好的清除ROS活性及较强的抗氧化能力,在化妆品领域有潜在应用前景。     

脂质过氧化引起的DNA损伤研究进展

脂质过氧化可以引起各种碱基损伤、DNA链断裂和各种荧光产物生成,并对DNA分子鸟嘌呤碱基具有选择性损伤.过渡金属离子可以明显加深脂质过氧化对DNA的损伤程度.多种抗氧化剂、活性氧自由基清除剂对脂质过氧化引起的DNA损伤有一定程度的保护作用.具有致突、致癌作用的8－羟基鸟嘌呤已经观察到．脂质过氧化的致突变、致癌变作用机制引起了人们的极大兴趣．     

DNA Binding,Antioxidant Activity,and DNA Damage Protection of Chiral Macrocyclic Mn(III) Salen Complexes

We are reporting the synthesis, characterization, and calf thymus DNA binding studies of novel chiral macrocyclic Mn(III) salen complexes S‐1 , R‐1 , S‐2 , and R‐2 . These chiral complexes showed ability to bind with DNA, where complex S‐1 exhibits the highest DNA binding constant 1.20 × 10⁶ M^?1. All the compounds were screened for superoxide and hydroxyl radical scavenging activities; among them, complex S‐1 exhibited significant activity with IC₅₀ 1.36 and 2.37 μM, respectively. Further, comet assay was used to evaluate the DNA damage protection in white blood cells against the reactive oxygen species wherein complex S‐1 was found effective in protecting the hydroxyl radicals mediated plasmid and white blood cells DNA damage. Chirality 24:1063–1073, 2012.© 2012 Wiley Periodicals, Inc.     

Sirtuins家族在DNA损伤修复中的作用

Sirtuins家族蛋白是一类依赖NAD的去乙酰化酶,属于第Ш类去乙酰化酶(HDACs),哺乳动物Sirtuins家族成员共有7个(SIRT1-7),其主要具有去乙酰化酶的活性,可以使多种蛋白发生去乙酰化,进而参与DNA的损伤修复、基因的转录调控、细胞凋亡、代谢及衰老等诸多生物进程。本文主要对Sirtuins家族在DNA损伤修复中的作用及其相关机制进行阐述。