老鼠Tet1蛋白能氧化单链脱氧核糖核酸上的5-甲基胞嘧啶到5-羟甲基胞嘧啶和5-羧基胞嘧啶(英文)

5-甲基胞嘧啶在哺乳动物细胞中具有广泛的作用.而它被双脱氧家族Tet蛋白氧化所得的产物5-羟甲基胞嘧啶、5-醛基胞嘧啶和5-羧基胞嘧啶也被证明在细胞发育和5-甲基胞嘧啶动态平衡调控中具有关键的作用.已有的研究结果表明,Tet蛋白能够识别双链DNA上的5-甲基胞嘧啶,并将其氧化成5-羟甲基胞嘧啶、5-醛基胞嘧啶和5-羧基胞嘧啶.我们通过质谱仪检测发现,老鼠Tet1蛋白的DNA结合结构域还能识别和氧化单链DNA上的5-甲基胞嘧啶.这一发现暗示我们,Tet蛋白家族不但具有已经发现的氧化双链DNA上5-甲基胞嘧啶的功能,还有可能作用于DNA的复制及转录,甚至具有氧化单链RNA上对应的甲基修饰碱基的能力.     

溴代胞嘧啶分子内三螺旋DNA的理论研究

运用分子力学方法研究了 3个含溴代胞嘧啶的分子内三螺旋及相应的未溴代分子内三螺旋DNA .理论结果表明 :当分子内三螺旋Hoogsteen嘧啶链的胞嘧啶被溴代胞嘧啶取代后其稳定性最强 ;而当分子内三螺旋 2条嘧啶链的胞嘧啶同时被溴代后 ,其稳定性最差 .从分子力学计算得到 ,4种分子内三螺旋的碱基以C3′ endo糖环构象为主 .在三螺旋YTCY( 2条嘧啶链中的胞嘧啶均被溴代胞嘧啶取代 )中 ,也含有一定数量的C2′ endo糖环构象 .因此 ,分子内三螺旋YTCY是A DNA和B DNA的综合体 ,而其他 3种三螺旋则是典型的A DNA .     

5-取代嘧啶核苷类似物的设计、合成及生物活性研究进展

综述了近年来有关新型5-取代嘧啶核苷类似物作为潜在生物活性先导化合物的设计、合成及生物活性研究进展.该类先导物的设计与合成主要通过以下两个方面来完成:一是在嘧啶碱基的5位上引入不同的取代基;二是在嘧啶碱基5位上引入取代基的同时对核苷中的糖基进行修饰,以得到新的衍生物.本文从以上两个方面出发,介绍了近年来这一研究领域所取得的主要进展.     

γ-核酸碱基取代丙基甲基二乙氧基硅烷的合成及其聚合反应研究

本文以Υ-溴丙基甲基二乙氧基硅烷分别与尿嘧啶、胸腺嘧啶、腺嘌呤和5-氟尿嘧啶进行烷基化反应,制得了四种含有核酸碱基或5-氟尿嘧啶的新型有机硅单体。通过它们的缩聚反应,合成了六种侧基含核酸碱基或5-氟尿嘧啶的聚硅氧烷。     

表观遗传修饰——5-羟甲基胞嘧啶检测的研究进展

表观遗传修饰是指在不改变DNA序列的情况下, 基因表达发生的可遗传变化. 5-羟甲基胞嘧啶(5-hydroxymethylcytosine, 5hmC)是继5-甲基胞嘧啶(5-methylcytosine, 5mC)后发现的表观遗传修饰, 被称为“第六种碱基”. 5hmC广泛分布于哺乳动物的组织和细胞中, 它的异常表达与肿瘤发生、发育性疾病和神经系统疾病密切相关. 由于5hmC的结构与5mC相似, 并且其丰度远远低于5mC, 传统方法难以实现对5hmC的准确和灵敏检测. 近年来, 科学家们结合新的修饰方法和信号放大策略, 发展了一系列超灵敏检测5hmC的新方法, 包括液相色谱串联质谱法、荧光法、电化学法、光电化学法、单碱基分辨率的测序法和单分子检测技术. 这些方法各自具有其独特优势, 有力推动了表观遗传学的发展. 本综述总结了5hmC检测方法的最新研究进展, 并对其面临的挑战和发展趋势做了展望.     

液相色谱-串联质谱法同时测定生物组织全基因组DNA甲基化和羟甲基化水平

测定全基因组DNA甲基化和羟甲基化水平对于研究环境污染物暴露的影响及致病机理具有重要的作用。本文建立了液相色谱-串联质谱（LC-MS/MS）同时测定动物组织中全基因组DNA甲基化和羟甲基化水平的方法。从动物组织样品中提取DNA,并将其酶解成单核苷,利用液相色谱-串联质谱法测定5-甲基胞嘧啶核苷、5-羟甲基胞嘧啶核苷和鸟嘌呤核苷的含量,计算全基因组DNA甲基化率和羟甲基化率。利用该方法研究了砷暴露对大鼠肝脏和小脑全基因组DNA甲基化和羟甲基化水平的影响,得到了砷影响DNA甲基化及羟甲基化的初步数据。该方法具有良好的重现性、灵敏度和稳定性,可以同时检测差异较大的DNA甲基化和羟甲基化水平。为同时研究DNA甲基化和羟甲基化水平提供了有力的支持。     

液相色谱-串联质谱分析组织中基因组脱氧核糖核酸羟甲基胞嘧啶水平

5-羟甲基胞嘧啶通过阻止脱氧核糖核酸(DNA)甲基化转移酶1(DMNT1)甲基化胞嘧啶来影响DNA甲基化的程度。本文建立了液相色谱-串联质谱(LC-MS/MS)测定组织中全基因组5-羟甲基胞嘧啶水平的方法。采用苯酚-氯仿提取组织DNA,提取的DNA用88%甲酸在140 ℃下裂解,DNA裂解液加入同位素胞嘧啶作内标,经N2吹干后,加乙腈-水(9:1, v/v)溶解,用LC-MS/MS检测5-羟甲基胞嘧啶的含量,并计算全基因组中5-羟甲基胞嘧啶的水平。结果表明,5-羟甲基胞嘧啶的线性范围为0.1～30 ng/mL,相关系数为0.9969,检出限(信噪比为3计)和定量限(信噪比为10计)分别为0.057 ng/mL和0.090 ng/mL;日内相对标准偏差和日间相对标准偏差分别为5.13%和6.24%;加标回收率为90.24%～97.53%。用该方法检测了大鼠大脑组织DNA羟甲基化水平,平均结果为0.66%。该方法简便,重现性好,灵敏度较高,能满足全基因组5-羟甲基胞嘧啶定量检测的要求。     

二乙二醇二甲基丙烯酸酯交联的胸腺嘧啶分子印迹聚合物纤维素支撑膜对核酸碱基分子选择性研究

使用光度分析法探讨了功能单体9-乙烯腺嘌呤和模板分子胸腺嘧啶在甲醇中的结合作用。在此基础上，以二乙二醇二甲基丙烯酸酯为交联剂，纤维素膜为支持体，制备了分子印迹聚合物膜。通过扫描电镜观察并比较了分子印迹聚合物膜、非分子印迹聚合物膜和纤维素载体膜通道的差异，以胸腺嘧啶、尿嘧啶、胞嘧啶、鸟嘌呤和腺嘌呤为底物，评价了印迹聚合物膜的渗透选择性。结果表明分子印迹膜对模板分子胸腺嘧啶及结构类似物尿嘧啶呈现高的选择性，与用乙二醇二甲基丙烯酸酯为交联剂制备的分子印迹膜比较，也呈现高的扩散速率和选择性，因此该膜对生物样品DNA完全水解产物胸腺嘧啶和RNA水解产物尿嘧啶的识别展现了良好的应用前景。     

核酸碱基与甘氨酸二肽间氢键作用的最佳位点

优化得到了碱基腺嘌呤、胸腺嘧啶、尿嘧啶、鸟嘌呤及胞嘧啶与甘氨酸二肽分子形成的28个氢键复合物的稳定结构并计算了结合能, 探讨了五种碱基与甘氨酸二肽分子间氢键作用的最佳位点. 本文研究发现: 每种碱基均可以通过不同位点与二肽分子形成氢键复合物, 腺嘌呤、胸腺嘧啶、尿嘧啶、鸟嘌呤及胞嘧啶分别最倾向使用A3、T1、U1、G3及C1位点与甘氨酸二肽分子形成氢键复合物; 碱基分子某位点的质子化反应焓变越负所形成的氢键复合物越稳定, 去质子化反应焓变越小所形成的氢键复合物越稳定; 由氢键复合物的结合能计算得到的稳定性次序与由碱基分子质子化和去质子化反应焓变推得的稳定性次序一致.     

DNA甲基化-非甲基化碱基间堆积作用的理论研究

运用二级Mфller-Plesset(MP2)理论方法和cc-pVDZ基组优化了6-甲基鸟嘌呤(O6-MethylG),4-甲基胸腺嘧啶(O4-MethylT)以及5-甲基胞嘧啶(C5-MethylC)与DNA碱基鸟嘌呤(G),腺嘌呤(A),胞嘧啶(C),胸腺嘧啶(T)之间的堆积构型.在MP2/aug-cc-pVXZ//MP2/cc-pVDZ(X=D,T)水平上,采用完全基组外推方法校正了堆积碱基对间的相互作用能,并用完全均衡校正法(CP)校正了基组重叠误差(BSSE).MP2计算结果表明,DNA碱基甲基化使得嘧啶-嘧啶、嘧啶-嘌呤堆积碱基间的平行旋转角发生明显改变,并使堆积碱基间的相互作用能增大.在MP2/cc-pVDZ计算级别上得到了各堆积碱基对的全电子波函数,并用分子中的原子理论(AIM)分析了堆积碱基对间的弱相互作用.AIM分析结果显示,甲基化增强了堆积碱基间的π-π作用,且甲基氢与相邻碱基间形成H2CH…X(X=O,N,CH3,NH2)等类型的氢键.甲基化损伤使碱基间重叠程度增大、π-π作用增强以及堆积碱基间形成多个氢键,是堆积作用能增加的主要原因.     

Quantification of 5-methylcytosine, 5-hydroxymethylcytosine and 5-carboxylcytosine from the blood of cancer patients by an enzyme-based immunoassay

Background

Genome-wide aberrations of the classic epigenetic modification 5-methylcytosine (5mC), considered the hallmark of gene silencing, has been implicated to play a pivotal role in mediating carcinogenic transformation of healthy cells. Recently, three epigenetic marks derived from enzymatic oxidization of 5mC namely 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), have been discovered in the mammalian genome. Growing evidence suggests that these novel bases possess unique regulatory functions and may play critical roles in carcinogenesis.

Methods

To provide a quantitative basis for these rare epigenetic marks, we have designed a biotin–avidin mediated enzyme-based immunoassay (EIA) and evaluated its performance in genomic DNA isolated from blood of patients diagnosed with metastatic forms of lung, pancreatic and bladder cancer, as well as healthy controls. The proposed EIA incorporates spatially optimized biotinylated antibody and a high degree of horseradish-peroxidase (HRP) labeled streptavidin, facilitating signal amplification and sensitive detection.

Results

We report that the percentages of 5mC, 5hmC and 5caC present in the genomic DNA of blood in healthy controls as 1.025 ± 0.081, 0.023 ± 0.006 and 0.001 ± 0.0002, respectively. We observed a significant (p < 0.05) decrease in the mean global percentage of 5hmC in blood of patients with malignant lung cancer (0.013 ± 0.003%) in comparison to healthy controls.

Conclusion

The precise biological roles of these epigenetic modifications in cancers are still unknown but in the past two years it has become evident that the global 5hmC content is drastically reduced in a variety of cancers. To the best of our knowledge, this is the first report of decreased 5hmC content in the blood of metastatic lung cancer patients and the clinical utility of this observation needs to be further validated in larger sample datasets.     

双核Pt(Ⅳ)配合物与Guanosine-5''''-Monophosphate和Glutathione反应的核磁共振光谱研究

合成了一个新型的双核Pt(Ⅳ)配合物{[cis-Pt(NH3)2Cl(OH)2]2(4,4'-methylenedianiline)}(NO3)2(化合物1)及相应的 15N标记化合物{[cis-Pt(15NH3)2Cl(OH)2]2(4,4'-methylenedianiline)}(NO3)2(化合物15N-1).利用1H NMR和ESMS进行了结构表征,化合物15N-1的2D[1H,15N]HSQC NMR发现,该化合物在水溶液中存在同分异构体.2D[1H,15N]HSQC NMR技术跟踪了化合物15N-1与Guanosine-5'-Monophosphate(5'-GMP)和Glutathione(GSH)的反应.结果显示,5'-GMP能在0.5 h内将化合物1还原,而GSH在6 h以后才能够部分的将化合物1还原.化合物1所表现出来的反应性能将有利于提高其治疗效果和降低毒副作用.     

Fructose dehydration to 5HMF in a green self-catalysed DES composed of N,N-diethylethanolammonium chloride and p-toluenesulfonic acid monohydrate (p-TSA)

Due to the increasing concerns about the availability and accessibility of fossil fuel reserves, and the subsequent effect of using them on climate change, production of green energy has recently become a hot area of interest in the research field. As a renewable energy source, biomass conversion to biofuels has shown a great potential towards green fuel production; particularly fructose conversion to 5-hydroxymethylfurfural (5HMF) as a building block material and source of green fuels and other high value chemicals.Herein, we investigate fructose dehydration to 5-hydroxymethylfurfural (5HMF) as a green fuel precursor, using a green self-catalysed environmentally friendly Deep Eutectic Solvent (DES), composed of inexpensive N,N-diethylethanolammonium chloride as organic salt and p-toluenesulfonic acid monohydrate (p-TSA) as a hydrogen bond donor (HBD) and novel medium for the fructose dehydration reaction.The advantage of using this DES is its ability to act as a solvent and catalyst simultaneously. It has shown to actively catalyse the dehydration reaction of fructose under moderate reaction conditions with a high 5HMF yield of 84.8% at a reaction temperature of 80 °C, reaction time of 1 h, DES mixing ratio of 1:0.5 salt to p-TSA (w/w), and initial fructose ratio of 5.     

The facile synthetic method of 5-hydroxymethyluracil

One practical and industrial procedure for preparation of 5-hydroxymethyluracil has been developed. This method has the advantage of facile operation, low cost and stable yield.     

Thermal analysis of laser-controlled 5-aminotetrazole propellant

Resorting to the continuous assistance of external laser radiation, laser augmented chemical propulsion (LACP) reveals superior combustion performance. However, the corresponding combustion theories have still to be ascertained for this innovative propulsion technique. Thermal characteristics of 5-aminotetrazole (5-ATZ) propellant are studied for LACP. In nitrogen environment, unique structures of burning surface are formed under laser ablation in bare samples of 5-ATZ propellant pressed powders. Resorting to an outer tube, a thermocouple is inserted in 5-ATZ propellant samples to obtain the temperature trace during combustion. In analogy with the burning surface structure reconstructed by micro computed tomography (MicroCT), the temperatures values of three significant regions were obtained. The experimental dependences of the measured temperatures on environment pressure and laser power are discussed.     

五价钒电解液稳定性研究

V 5的稳定性是指在溶液中不出现V2O5轻微沉淀的时间。钒电池是一种反复可充的高效储能新型无污染化学电池[1],与其它电池相比,钒电池具有不可替代的优势[2,3]。该电池活性物质是溶解在强酸溶液中的钒的氧化物或化合物。电解液中钒离子的浓度大小和电解液的多寡决定了电池的容量     

STUDY ON THE MECHANISM OF INTERFERON ACTION (Ⅺ)——EFFECT OF pppA2''''p5''''A2''''p5''''A ON THE LEVEL OF cAMP AND cGMP IN MACROPHAGES

In this paper, the increase of cellular cAMP and cGMP levels in macrophages induced bypppA2'p5'A2'p5'A (briefly 2'-5'P_3A_3) is first reported. The optimal concentration of 2'-5'P_3A_3 for the elevation of cellular cGMP to the highest level is 10~(-7)-10~(-6)mol/L, while thatfor cAMP is 10~(-7)mol/L. The time for cGMP to reach its peak value is 15 min and that forcAMP is 2 h, when the cells are treated with 2'-5' P_3A_3 at 10~(-7)mol/L, which is the optimalconcentration for developing biological effect of macrophages (phagocytosis). These resultssuggest that cGMP and cAMP may be related to, or may be the mediators for, 2'-5'P_3A_3action.