Chemistry and Biology of Malondialdehyde DNA Adducts

描述（由申请人提供）：丙二醛（MDA）和其他氧化应激醛类产物是一类重要的内源性DMA损伤剂，普遍存在于人体中。这些双功能亲电试剂产生脱氧核苷碱基的外环加合物，阻止沃森-克里克碱基配对，阻断DNA复制并诱导突变。我们的实验室已经确定了嘧啶嘌呤酮加合物脱氧鸟苷（M1dG）的化学和生物学性质，这是DNA被丙二醛和基丙烯损伤的主要产物。双链DNA催化M1dG与其开环衍生物N2-（3-氧丙烯基）-脱氧鸟苷（OPdG）之间的动态平衡。与M1dG相比，OPdG对DNA复制的阻断作用更小，与稳定的M1dG外环类似物相比，OPdG的致突变性更小。我们最近发现M1dG在大肠杆菌和哺乳动物细胞中诱导重复d(CG)n序列的序列依赖移码突变。我们假设M1dG诱导移码突变的能力是由于它转化为OPdG，我们建议通过直接比较M1dG和OPdG的一系列结构类似物诱导d(CG)n序列移码突变的能力来验证这一假设。我们最近开发了一种在寡核苷酸中合成MDA-DNA加合物的方法，并打算用它来探索另一种主要的DNA加合物N6-（3-氧丙烯基）-脱氧腺苷（OPdA）的化学和生物学性质。我们将确定哺乳动物细胞中OPdA的突变谱，并验证OPdA和氧化应激的其他脱氧腺苷产物诱导重复dAn序列移码突变的假设。这种类型的突变通常在导致人类癌症的关键生长调节基因中观察到，其发生与氧化应激有关。据报道，在暴露于氧化剂的细胞中存在mda诱导的dna -蛋白质交联，但这些交联的身份及其形成的化学过程却非常不明确。我们拟探索M1dG、OPdG和OPdA与氨基酸、多肽和蛋白质的化学反应。将特别关注与dma结合蛋白的结合物的鉴定，如限制性内切酶、核苷酸切除修复酶和组蛋白。这些实验将为评估mda诱导的dna -蛋白交联是细胞氧化损伤的重要产物这一假设提供关键的化学信息。

DESCRIPTION (provided by applicant): Malondialdehyde (MDA) and other aldehydic products of oxidative stress represent an important class of endogenous DMA damaging agents universally generated in human beings. These bifunctional electrophiles generate exocyclic adducts to deoxynucleoside bases that prevent Watson-Crick base pairing, block DNA replication, and induce mutations. Our laboratory has defined the chemistry and biology of the pyrimidopurinone adduct to deoxyguanosine, M1dG, which is the major product of DNA damage by MDA and base propenals. Duplex DNA catalyzes a dynamic equilibrium between M1dG and its ring-opened derivative, N2-(3-oxopropenyl)-deoxyguanosine (OPdG). OPdG is less blocking than M1dG to DNA replication and less mutagenic than stable exocyclic analogs of M1dG . We recently discovered that M1dG induces sequence-dependent frameshift mutations in reiterated d(CG)n sequences in E. coli and mammalian cells. We hypothesize that the ability of M1dG to induce frameshift mutations is due to its conversion to OPdG and we propose to test this hypothesis by directly comparing the ability of a series of structural analogs of M1dG and OPdG to induce frameshift mutations in d(CG)n sequences. We recently developed a general synthesis of MDA-DNA adducts in olignucleotides and we propose to use it to explore the chemistry and biology of the other major DNA adduct, N6-(3-oxopropenyl)-deoxyadenosine (OPdA). We will determine the mutation spectrum of OPdA in mammalian cells and we will test the hypothesis that OPdA and other deoxyadenosine products of oxidative stress induce frameshift mutations in reiterated dAn sequences. This type of mutation is commonly observed in critical growth regulating genes that contribute to human cancers and its occurrence has been associated with oxidative stress. MDA-induced DNA-protein cross-links have been reported to exist in cells exposed to oxidants but the identity of these cross-links and the chemistry of their formation is very poorly defined. We propose to explore the chemistry of reaction of M1dG, OPdG, and OPdA with amino acids, peptides, and proteins. Particular attention will be given to the identification of conjugates with DMA-binding proteins such as restriction endonucleases, nucleotide excision repair enzymes, and histones. These experiments will provide critical chemical information with which to evaluate the hypothesis that MDA-induced DNA-protein cross-links are important products of oxidative damage to cells.