MOLECULAR PHARMACOLOGY OF TUMOR AND VIRUS INHIBITORS

肿瘤和病毒抑制剂的分子药理学

• 批准号: 2007200
• 负责人: Arthur Patrick Grollman
• 金额: $ 39.87万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 1977
• 资助国家: 美国
• 起止时间: 1977-01-01 至 1998-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2007200
• 关键词: DNA damage; DNA directed DNA polymerase; DNA repair; antiviral agents; carcinogenesis; carcinogenesis inhibitor; chemical binding; endonuclease; enzyme mechanism; enzyme substrate; frameshift mutation; gene mutation; molecular genetics; neoplasm /cancer genetics; neoplasm /cancer pharmacology; nucleic acid sequence; nucleic acid structure; oligonucleotides; site directed mutagenesis

This research focusses on the biological consequences of DNA damage with the overall goal of elucidating primary events in carcinogenesis. A principal theme is to establish relationships between the structure of damaged DNA and the functions of enzymes involved in DNA replication and repair. Novel experimental systems have been developed for this purpose. Site specific mutagenesis involves a strategy in which a shuttle plasmid vector, containing a single defined lesion, is allowed to replicate in mammalian cells or bacteria. The position of mutations induced is established by DNA sequence analysis. Primer-extension reactions, catalyzed by DNA polymerase, coupled with steady-state kinetic analysis, are used to explore translesional synthesis and mutagenic events in vitro. Our specific aims are (a) to establish models for frameshift mutagenesis in terms of misaligned DNA templates and kinetics governing translesional synthesis; (b) to elucidate the molecular basis underlying sequence context effects on base substitutions and deletions; (c) to understand the role of SOS functions in translesional synthesis; (d) to discover pathways by which mutations are generated during repair of bistrand abasic sites in DNA; (e) to demonstrates differences between DNA polymerases in their abilities to generate mutations arising from DNA damage; (f) to develop in vitro assays that predict mutagenic specificity for defined DNA lesions in vivo; (g) to explore mechanisms by which DNA damage enhances the frequency of homologous recombination in mammalian cells and bacteria; and (h) to establish the solution structure of misaligned intermediates formed during deletion mutagenesis. Additional studies are designed (a) to determined the substrate specificity of Fpg protein; (b) to establish the role of the N-terminus in the catalytic function of this enzyme; (c) to reveal the structural basis for binding of the zinc finger domain oxidatively-damaged DNA; (d) to elucidate a catalytic mechanism for DNA glycosylate activity; (e) to detect functional groups on Fpg protein and its substrates that facilitate "recognition" of oxidative damage; (f) to establish the structure of complexes formed between Fpg protein or adenine DNA glycosylate and analogs of their DNA substrates; and (g) to study substrate binding and mechanism of action of selected AP endonucleases, and (h) to quantify the contribution of Fpg protein to DNA repair in cells.

这项研究的重点是DNA损伤的生物学后果 阐明癌症发生中的主要事件的总体目标。一个 主要的主题是建立结构之间的关系 DNA损伤和参与DNA复制的酶的功能和 修理。为此目的开发了新的实验系统。 定点突变涉及一种策略，在该策略中，穿梭质粒 包含单个已定义病变的载体被允许在 哺乳动物细胞或细菌。诱发突变的位置是 通过DNA序列分析建立。引子延伸反应， 由DNA聚合酶催化，结合稳态动力学分析， 用来探索变态合成和诱变事件 体外培养。 我们的具体目标是：(A)建立移码突变模型 在错位的DNA模板和控制转位的动力学方面 合成；(B)阐明序列的分子基础 上下文对碱基替换和删除的影响；(C)了解 SOS功能在翻译合成中的作用；(D)发现 双链修复过程中产生突变的途径 DNA中的基本位置；(E)显示DNA之间的差异 聚合酶产生DNA突变的能力 损害；(F)开发预测突变特异性的体外试验 用于体内明确的DNA损伤；(G)探索DNA 损伤增加哺乳动物中同源重组的频率 细胞和细菌；及(H)建立其溶液结构 在缺失突变过程中形成的错位中间产物。 设计了额外的研究(A)以确定底物 FpG蛋白的特异性；(B)确定N末端的作用 该酶的催化功能；(C)揭示其结构 锌指结构域结合氧化损伤的DNA的基础；(D) 阐明DNA糖基化活性的催化机制；(E) 检测FPG蛋白及其底物上的官能团 促进对氧化损害的“认识”；(F)建立 FPG蛋白与腺嘌呤DNA形成的复合体的结构 糖基化及其DNA底物的类似物；及(G)研究 选定的AP内切酶的底物结合和作用机制， 和(H)量化FPG蛋白在DNA修复中的作用 细胞。