博安霉素与DNA相互作用的分子机理研究

通过光谱分析、粘度测定及~１ＨＮＭＲ研究证实：博安霉素（ＢｌｅｏｍｙｃｉｎＡ_６,ＢＬＭＡ_６）是通过双噻唑基嵌插入碱基对之间与ＤＮＡ结合的。同时测定了ＢＬＭＡ_６与ＤＮＡ的结合常数、结合位点数并与博莱霉素Ａ_２（ＢＬＭＡ_２）、Ａ_５（ＢＬＭＡ_５）进行了比较,证实了末端胺基对ＢＬＭＡ_６与ＤＮＡ结合的贡献,琼脂糖凝胶电泳对ＢＬＭＡ_６及其Ｃｕ（Ⅱ）、Ｆｅ（Ⅱ）络合物断裂ＤＮＡ的研究表明,在ＤＮＡ断裂中某种氧自由基的存在及金属螯合部位与ＤＮＡ嵌插部位之间的相互影响,对于ＢＬＭＡ_６及同系物对小鼠肺毒性的差异与不同尾链结构的关系进行了探讨。     

经野生稻总DNA处理的栽培稻离体胚长成植株后的某些性状变化(简报)

栽培稻离体胚（幼胚、萌动胚）以野生稻总ＤＮＡ浸泡或滴加处理后,经组织培养长成的完整植株进行盆栽的结果显示,当代与后代植株中大多数无变异,仅１株产生芒,２株抗稻瘟病。有芒种子再次种植于人工气候室内,其Ｄ_１、Ｄ_２和Ｄ_３代种子仍有芒;抗稻瘟病植株的种子收获后再次播种,后代仍有抗稻瘟病特性。     

双歧杆菌总DNA对小鼠IL—2,NK活性影响的研究

目的 从双歧杆菌、大肠杆菌提取ＤＮＡ,用ＤＮＡ免疫小鼠,观察免疫功能的变化,探讨双歧杆菌ＤＮＡ对小鼠免疫功能的影响,并作对比研究。方法 肌肉注射提取的双歧杆菌ＤＮＡ、大肠杆菌ＤＮＡ,颈椎处死后,检测脾细胞的免疫功能,同时提取ＩＥＬ细胞与ＤＮＡ共孵育,检测它对ＩＥＬ细胞的激活情况及细胞因子产生情况,以自然杀伤细胞（ＮＫ）活性,白细胞介素２（ＩＬ－２）产生能力为指标,测定小鼠上述各项指标变化。结果 双歧杆菌ＤＮＡ、大肠杆菌ＤＮＡ肌肉注射后,小鼠以上两项指标与相应对照组相比较均明显提高（Ｐ〈０．０５）。双歧杆菌ＤＮＡ提高小鼠ＮＫ活性与ＩＬ－２水平程度大于大肠杆菌ＤＮＡ的作用（Ｐ〈０．０１）。结论 双歧杆菌ＤＮＡ可快速激活ＮＫ活性,提高体内ＩＬ－２水平。其效能优于大肠杆菌ＤＮＡ。     

细胞松驰素B促微丝解聚对DNA合成的作用

利用微丝（ｍｉｃｒｏｆｉｌａｍｅｎｔ,ＭＦ）解聚药物细胞松驰素Ｂ（ｃｙｔｏｃｈａｌａｓｉｎＢ,ＣＢ）处理Ｇ_０期小鼠Ｃ_３Ｈ_（１０）Ｔ_（１／２）成纤维细胞,对Ｇ_０至Ｓ期ＤＮＡ合成,胸腺嘧啶核苷激酶（ｔｈｙｍｉｄｉｎｅｋｉｎａｓｅ,ＴＫ）活性、ＴＫ基因表达、钙调素（ｃａｌｍｏｄｕｌｉｎ,ＣａＭ）水平和一些细胞周期早期基因的表达进行了观察,Ｇ_０期细胞经３ｍｇ／ＬＣＢ处理２ｈ,促ＭＦ解聚增强了血清对Ｓ期细胞ＴＫ活性、ＴＫ基因表达和ＤＮＡ合成的刺激作用,并促进细胞提前进入Ｓ期．血清刺激Ｇ_０期细胞进入晚Ｇ_１期和Ｓ期时,ＣａＭ水平明显升高,而ＣＢ预处理则使ＣａＭ含量进一步增加,特别是ＣＢ处理促使Ｓ期ＣａＭ增加向核内转移．ＣＢ处理明显增强血清对ｃ－ｊｕｎ、ｃ－ｆｏｓ和ｃ－ｍｙｃ基因表达的刺激作用,而ＰＫＣ抑制剂Ｈ_７则抑制ＣＢ处理对这些基因转录的刺激作用,说明ＣＢ使Ｇ_０期细胞ＭＦ解聚刺激ｃ－ｊｕｎ、ｃ－ｆｏｓ和ｃ－ｍｙｃ的转录活性与ＰＫＣ的作用有关．结果表明Ｇ_０至Ｓ期早期ＭＦ的重组可促进细胞进入Ｓ期,增强ＤＮＡ合成．     

H_2O_2－Fe~（3＋）所致人淋巴细胞DNA双链断裂损伤

采用脉冲电场凝胶电泳法检测Ｈ２Ｏ２－Ｆｅ（３＋）体系产生的ＯＨ对人淋巴细胞ＤＮＡ的双链断裂损伤．Ｈ２Ｏ２－Ｆｅ（３＋）浓度与ＤＮＡ双链断裂呈明显量效关系;随ＯＨ作用时间延长,细胞ＤＮＡ双链断裂加重;过氧化氢酶对ＯＨ损伤有明显抑制作用．脉冲电场凝胶电泳法可检测到的Ｈ２Ｏ２和ＦｅＣｌ３引起细胞ＤＮＡ双链断裂的最低浓度为０．３ｍｍｏｌ／Ｌ和６μｍｏｌ／Ｌ．     

丙型肝炎病毒E2基因DNA免疫实验动物研究

将编码丙型肝炎病毒（ＨＣＶ）Ｅ２蛋白４１７～７５０位氨基酸的ＤＮＡ片段　克隆到真核表达载体ｐｃＤＮＡ　３．１（－）中的ＣＭＶ　ＩＥ启动子下游,构建成ＨＣＶ　Ｅ２重组真核表达质粒　ｐｃＥ２。ＥＬＩＳＡ法检测ｐｃＥ２　ＤＮＡ免疫兔血清中的Ｅ２抗体变化和维持规律,结果显示免疫２０ｄ已有　抗体产生,３０ｄ后开始进入高峰,４０ｄ时达到最高值,至第９０ｄ抗体水平保持平稳,抗体滴度　达到１∶１６００左右。流式细胞计数仪（ＦＡＣＳ）检测ｐｃＥ２　ＤＮＡ免疫鼠ＣＤ４＋、ＣＤ８＋Ｔ淋巴细胞变　化情况,与注射空载体ｐＣＤＮＡ３．１（－）的阴性鼠相比,ＣＤ４＋淋巴细胞水平略有上升,ＣＤ８＋　细胞水平有较大升高,增幅达３５．４６％。免疫组化检测结果显示注射ｐｃＥ２的小鼠组织中有明显　的阳性着色,而注射ｐｃＤＮＡ３．１（－）的对照组小鼠免疫组化结果为阴性。以上结果表明：ｐｃＥ２　在实验动物内表达出的ＨＣＶ　Ｅ２蛋白可以引起免疫动物的体液免疫应答和细胞免疫应答,尤其　是ＭＨＣ－１限制性杀伤性ＣＤ８＋Ｔ淋巴细胞水平的提高对清除　病毒是十分有利的,因此ＨＣＶ　Ｅ２　ＤＮＡ免疫有可能成为预防和治疗ＨＣＶ感染的一条新途径。     

前列腺素F2α对大鼠肾小球系膜细胞DNA合成的影响

本实验应用（＾３Ｈ）胸腺嘧啶核苷（＾３Ｈ－ＴｄＲ）掺入法测定前列腺素Ｆ２α（ＰＧＦ２α）对大鼠肾小球系膜细胞ＤＮＡ合成的作用,测定系膜细胞合成的二脂酰甘油（ＤＡＧ）及磷酸肌醇（ＩＰ）。结果表明,ＰＧＦ２α促进系膜细胞的ＤＮＡ合成、ＤＡＧ及ＩＰ的生成。提示ＰＧＦ２α使系膜细胞的磷脂酶Ｃ活化,产生ＩＰ及ＤＡＧ,激活蛋白激酶Ｃ,从而促进ＤＮＡ合成及细胞增殖。     

DNA损伤与修复

《ＤＮＡ损伤与修复,（第２卷）：高级真核生物的ＤＮＡ修复》（ＤＮＡＤａｍａｇｅａｎｄＲｅｐａｉｒ,ＶｏｌｕｍｅＩＩ：ＤＮＡＲｅｐａｉｒｉｎＨｉｇｈｅｒＥｕｋａｒｙｏｔｅｓ）由ＪａｃＡ．Ｎｉｃｋｏｌｏｆｆ和ＭｅｒｌＦ．Ｈｏｅｋｓｔｒａ编著,１９９８年ＨｕｍａｎａＰｒｅｓｓ出版,６３９页。ＤＮＡ损伤与修复提供了在广泛多样化生物体范围内ＤＮＡ修复主要方面的重要和简洁评论。第２卷：高级真核生物的ＤＮＡ修复集中于哺乳动物系统、人体遗传学疾病和癌症,强调最近５年发展的重要过程。主要题目：紫外线和Ｘ线的修复,…     

博安霉素与DNA相互作用的分子机理研究

通过光谱分析,粘度测定及＾１ＨＮＭＲ研究证实,博安霉素（ＢｌｅｏｍｙｃｉｎＡ６,ＢＬＭＡ６）是通过双噻唑其嵌插入碱基对之间与ＤＮＡ结合,同时测定了ＢＬＭＡ６与ＤＮＡ的结合常数,结合位点数并与博霉素Ａ２（ＢＬＭＡ２）,Ａ５（ＢＬＭＡ５）进行了比较,证实了末端胺基对ＢＬＭＡ６与ＤＮＡ结合的贡献,琼脂糖凝胶电泳对ＢＬＭＡ６及其Ｃｕ（Ⅱ）,Ｆｅ（Ⅱ）络合物断裂ＤＮＡ的研究表明,在ＤＮＡ断裂中某种氧自由基的     

质粒DNA增强大鼠缺血骨骼肌肌浆网钙转运功能

在大鼠肢体缺轿模型上观察质粒ｐｃＤＮＡ３对缺血骨骼肌肌浆网（ＳＲ）Ｃａ＾２＋转运的影响。结果显示,骨骼肌缺血时ＳＲＣａ＾２＋转运（Ｃａ＾２＋摄入与释放）较非缺血肌肉增强,而质粒ｐｃＤＮＡ３与ＳＲ上ＤＮＡ结合蛋白结合之后,可进一步增强缺备骨骼肌ＳＲＣａ＾２＋摄入（Ｐ＜０．０１）及释放速率（Ｐ＜０．０５）。提示质粒ＤＮＡ对正常及缺血大鼠骨骼肌的ＳＲＣａ＾２＋转运能力均有影响,其临床病理生理意义值得进一     

Synthesis and properties of 2'-O-methyl-2-thiouridine and oligoribonucleotides containing 2'-O-methyl-2-thiouridine

A new method for the synthesis of 2'-O-methyl-2-thiouridine (s2Um) found in thermophilic bacterial tRNA was developed. Structural properties of s2Um and s2Um(p)U were studied by using 1H NMR spectroscopy. A modified nonaribonucleotide (RNA*: 5'-CGUUs2UmUUGC-3') was synthesized to study the base-recognition ability of s2Um in formation of RNA-RNA and RNA DNA duplexes. The UV melting experiments revealed that RNA*-RNA and RNA*-DNA duplexes having an s2U-A base pair are more stable than those having a U-A base pair. On the contrary, the thermal stability of RNA*-RNA and RNA*-DNA duplexes having an s2U-G wobble base pair was much lower than that of the unmodified duplexes having a natural U-G base pair. It is concluded that s2Um has higher selectivity toward A over G than unmodified U.     

Synthesis and physicochemical properties of 2'-deoxy-2',2'-difluoro-beta-D-ribofuranosyl and 2'-deoxy-2',2'-difluoro-alpha-D-ribofuranosyl oligonucleotides

We present procedures for nucleoside and oligonucleotide synthesis, binding affinity (Tm) and structural analysis (CD spectra) of 2'-deoxy-2',2'-difluoro-alpha-D-ribofuranosyl and 2'-deoxy-2',2'-difluoro-beta-D-ribofuranosyl oligothymidylates. Possible reasons for the thermal instability of duplexes formed between these compounds and RNA or DNA targets are discussed.     

Stereospecific 2'-amination and 2'-chlorination of adenosine by Actinomadura in the biosynthesis of 2'-amino-2'-deoxyadenosine and 2'-chloro-2'-deoxycoformycin

2'-Amino-2'-deoxyadenosine and 2'-chloro-2'-deoxycoformycin (2'-CldCF) are two nucleoside antibiotics produced by Actinomadura. The biosynthesis of these two nucleoside antibiotics has been studied by the addition of [U-14C]adenosine with or without unlabeled adenine to cultures of Actinomadura. By this experimental approach, it is possible to demonstrate that adenosine is the direct precursor for the biosynthesis of 2'-amino-2'-deoxyadenosine and 2'-CldCF. These conclusions are based on the observation that the percentage distribution of 14C in the aglyconic and pentofuranosyl moieties of 2'-amino-2'-deoxyadenosine and 2'-CldCF were similar to the distribution of 14C in the adenine and ribosyl moieties of the [U-14C]adenosine (i.e., 48:52) added to cultures of Actinomadura. Experimentally, the percentage distribution of 14C in the (i) adenine:2-amino-2-deoxy-beta-D-ribofuranose of 2'-amino-2'-deoxyadenosine is 51:49; (ii) 8-(R)-3,6,7,8-tetrahydroimidazo[4,5-d]-[1,3-diazepin-8-o1]:2 -chloro-2- beta-D-ribofuranose of 2'-CldCF is 45:55; and (iii) adenine:ribose of the adenosine isolated from the RNA of Actinomadura is 42:58. Further proof that adenosine is the direct precursor for the biosynthesis 2'-amino-2'-deoxyadenosine and 2'-CldCF was demonstrated by the addition of 75 mumol of unlabeled adenine together with [U-14C]adenosine to nucleoside-producing cultures of Actinomadura. The percentage distribution of 14C in the aglycon and the sugar moieties of 2'-amino-2'-deoxyadenosine and 2'-CldCF were 46:54 and 47:53, respectively; the percentage distribution of 14C in the adenine and ribose moieties of the adenosine isolated from the RNA of Actinomadura was 51:49. These data show that the hydroxyl on C-2' of the ribosyl moiety of adenosine undergoes a replacement by a 2'-amino or a 2'-chloro group to form 2'-amino-2'-deoxyadenosine or 2'-CldCF with retention of stereconfiguration at C-2'. Finally, Actinomadura can utilize inorganic chloride from the medium as demonstrated by the isolation of [36Cl]2'-CldCF following the addition of [36Cl]chloride to the culture medium. Mechanisms for the regioselective modification of the C-2' hydroxyl group and stereospecific insertion of the amino and chloro groups are discussed.     

Stereoselective Synthesis of 2-Amino-2-deoxy-d-arabinose and 2-Deoxy-d-ribose

An efficient method for the stereoselective synthesis of 2-amino-2-deoxy-d-arabinose and 2-deoxy-d-ribose is described.

The key step in this method was accomplished by the nucleophilic addition of methyl isocyanoacetate to 2,3-O-isopropylidene-d-glyceraldehyde with high erythro-selectivity (nearly 100%).

Subsequent intermolecular cyclization predominantly gave the desired oxazoline derivative (trans-form), in which two new chiral centers were formed. The oxazoline derivative was efficiently converted to both 2-amino-2-deoxy-d-arabinose and 2-deoxy-d-ribose.     

Characterization of alpha 2 beta 2 and alpha 2 forms of kinesin

Bovine brain kinesin separates into two components on sucrose density gradient centrifugation. The predominant component is a heterotetramer of two 120 kDa alpha subunits and two 64 kDa beta subunits with an sedimentation coefficient of 9.6 S and a low Vm rate of microtubule-stimulated ATPase of 1.3 +/- 0.5 sec-1 at 25 degrees, pH 7.0. The minor element is a homodimer of two alpha subunits without beta subunits with a sedimentation coefficient of 6.9 S and a higher Vm rate of microtubule-stimulated ATPase of 7.0 +/- 1.9 sec-1. Microtubules stimulate the rate of release of ADP from the active site of the tetramer, but the rate of release is not fast enough to account for the rate of steady state ATP hydrolysis. Further complexity is indicated by biphasic release kinetics. In spite of the large difference in Vm ATPase rate for the two species, both drive the sliding of sea urchin axonemes over glass surfaces at the same velocity.     

Complexes of arabinogalactan of Pereskia aculeata and Co2+, Cu2+, Mn2+, and Ni2+

The main interest in the biopolymer arabinogalactan is that it is edible. Complementing its high protein percentage, when complexed to essential metal ions, widens the use in food and pharmacology industries and technologies. The binding constants of Co2+, Cu2+, Mn2+ and Ni2+ with arabinogalactan, extracted from the leaves of Pereskia aculeata from Brazil were determined by potentiometric titrations and also the speciation according to pH values. The complexed species proposed by potentiometric titrations and the unique complexing ability of galacturonic acid groups towards Cu2+ and Ni2+ in the tridimensional web structure of arabinogalactan were confirmed by IR and EPR spectroscopies. The thermal stability of the complexed species also varied with the metal ion employed in the complexation when compared to the biopolymer alone. These complexes are new sources of additives for the food and pharmacology industries and carriers of essential metal ions to animal and vegetal biochemistry.