Sequence Effects of Arylamine-DNA Adducts: Repair and Replication

项目摘要/摘要 我们研究计划的长期目标是阐明dna的分子机制。 加合物诱导的化学致癌作用。芳香胺是众所周知的环境人类 致癌物质。特别是，芳胺-DNA加合物的形成已在各种人类中得到证实。 组织，并被认为会导致突变。我们之前已经证明了DNA中的芳胺加合物 以三种明确的形态存在：堆积型(S)、外部B型(B)和楔形(W)。这个 构象取决于致癌物部分在DNA分子中的位置，而 在生理条件下，各类型的种群比例是顺序相关的。在这 应用，我们假设芳胺诱导的修复和突变是构象(S/B/W) 具体的。我们提出了四个主要目标来帮助定义加合物构象并研究它们的特异性 对修复(初始损伤识别)和复制(复制分叉异质性)的影响。 具体地说，这些目标集中在：(1)人类核苷酸切除中的构象特异性修复 修复(NER)系统，(2)长程序列效应，(3)依赖于序列的热力学 滑移诱导的移码突变，以及(4)复制叉构象 异质性。我们将不仅使用现有的动态~(19)F核磁共振/CD、EMSA和荧光 光谱学，但也创新的基于芯片的表面等离子体共振(SPR)和差分 扫描量热(DSC)程序，创建了一套强大的生物物理方法。 成功完成提出的目标将有助于我们更好地掌握蛋白质-dna。 参与人类NER和跨病变合成的相互作用，这对 解析癌症病因学的分子细节。这些知识也将有助于 制定合理的预防和风险评估战略。

Project Summary/Abstract The long term goal of our research program is to elucidate the molecular mechanisms of DNA adduct-induced chemical carcinogenesis. Aromatic amines are well-known environmental human carcinogens. In particular, arylamine-DNA adduct formation has been confirmed in various human tissues and is believed to induce mutation. We have previously shown that arylamine adducts in DNA exist in three well-defined conformations: stacked (S), external B-type (B), and wedge (W). The conformation depends on the location of the carcinogen moiety in the DNA molecule, and the population ratios of the types under physiological conditions are sequence-dependent. In this application, we hypothesize that arylamine-induced repair and mutation is conformation (S/B/W) specific. We propose four major aims to help define adduct conformation and examine their specific effects on repair (initial damage recognition) and replication (replication fork heterogeneity). Specifically, these aims focus on: (1) conformation-specific repair in a human nucleotide excision repair (NER) system, (2) long-range sequence effects, (3) the thermodynamics of sequencedependent slippage-induced frameshift mutagenesis, and (4) replication fork conformational heterogeneity. We will employ not only existing dynamic 19F NMR/CD, EMSA and fluorescence spectroscopy, but also innovative chip-based surface plasma resonance (SPR) and differential scanning calorimetric (DSC) procedures, creating a powerful suite of biophysical methodologies. Successful completion of the proposed aims will help us gain a better grasp on the protein-DNA interactions involved in human NER and trans-lesion synthesis, which have important implications for resolving the molecular details of cancer etiology. Such knowledge will also be of help in the development of sensible prevention and risk assessment strategies.