DNA ALKYLATION DAMAGE, LESION STRUCTURE AND REPAIR

描述：（申请人的描述） 本申请涉及DNA 0-烷基化的结构生物学 损害 作为申请人长期目标的一部分， 分子识别受损的DNA，他建议确定 含有烷基损伤和碱基类似物的DNA双链体的溶液结构 目前用于化疗。 他将建立三维的 修复蛋白的结构：06-甲基鸟嘌呤甲基转移酶（Ogt）， 游离的和与酶抑制剂反应后的，以及受损的 DNA/蛋白质复合物。 烷基转移酶活性与 对甲基化剂诱导的突变的敏感性增加。 肿瘤 表达高水平烷基转移酶活性的细胞系是 抗治疗性甲基化和氯乙基化作用 剂. 此外，烷基转移酶抑制剂与 甲基化化疗药物克服了某些肿瘤的耐药性 细胞系与药物的关系 该项目产生的信息将 增加我们对化学诱变和DNA修复知识， 合理设计新型烷基转移酶抑制剂， 在癌症治疗中。 利用亚磷酰胺化学，他将合成 大量含有碱基类似物的寡脱氧核苷酸双链体 S6-硫代-2 '-脱氧鸟苷和S6-甲基-6-硫代-2'-脱氧鸟苷配对 与脱氧胞苷和胸苷相反。 它们的三维结构 将通过核磁共振波谱学和计算方法建立。 他将 利用重组DNA技术获得大量高纯度的OGT 天然丰富的蛋白质以及富含15 N和13 C。 他 溶液结构将通过使用多核-- 多维NMR光谱学和计算方法。 类似 方法将用于确定反应后蛋白质的结构 用酶抑制剂06-甲基鸟嘌呤和06-苄基鸟嘌呤。

DESCRIPTION: (Applicant's Description) This application deals with the structural biology of DNA 0-alkylation damage. As part of the applicant's long-term objectives of understanding the molecular recognition of damaged DNA, he proposes to determine the solution structure of DNA duplexes containing alkyl lesions and base analogs currently used in chemotherapy. He will establish the three-dimensional structure of the repair protein: 06-methylguanine methyltransferase (Ogt), free and after reaction with enzyme inhibitors, and of the damaged DNA/protein complex. Alkyl transferase activity has been correlated with increased sensitivity to mutations induced by methylating agents. Tumor cell lines expressing high levels of alkyl transferase activity are resistant to the action of therapeutic methylating and chloroethylating agents. In addition, alkyl transferase inhibitors used in combination with methylating chemotherapeutic agents overcome the resistance of some tumor cell lines to the drugs. The information resulting from this project will increase our knowledge of chemical mutagenesis and DNA repair and facilitate the rational design of new alkyl transferase inhibitors, which can be used in cancer therapy. Using phosphoramidite chemistry, he will synthesize large quantities of oligodeoxynucleotide duplexes containing the base analog S6-thio-2'-deoxyguanosine and S6-methyl-6-thio-2'-deoxyguanosine paired opposite to deoxycytidine and thymidine. Their three-dimensional structure will be established by NMR spectroscopy and computational methods. He will use recombinant DNA technology to obtain large quantities of highly pure Ogt protein in natural abundance as well as enriched with 15N and 13C. His solution structure will be determined by using multinuclear -- multidimensional NMR spectroscopy and computational methods. Similar methods will be used to determine the structure of protein after reaction with the enzyme inhibitors 06-methylguanine and 06-benzylguanine.