MASS SPECTROMETRY FOR DNA ADDUCTS STRUCTURES

DNA 加合物结构的质谱分析

• 批准号: 6344722
• 负责人: MICHAEL L GROSS
• 金额: $ 16.87万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2001-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6344722
• 关键词: DNA damage; adduct; animal genetic material tag; carbopolycyclic compound; chemical carcinogenesis; electrospray ionization mass spectrometry; estrogens; gas chromatography mass spectrometry; matrix assisted laser desorption ionization; nucleic acid structure; technology /technique development

Recently we have established that high-energy tandem mass spectrometry is sufficiently specific for determining structure of DNA adducts. When coupled with fast atom bombardment mass spectrometry, however, the detection limits and sensitivity are not adequate to support investigations of adducts in the femtomole range. To overcome this limitation, we propose to develop and compare three mass spectrometric approaches for high- sensitivity detection and structure determination of DNA adducts. Tandem foursector mass spectrometry with extended array detection has recently been coupled with electrospray ionization in our laboratory. Continued development is the highest priority because the product-ion spectra have sufficient specificity to distinguish isomeric adducts. Efforts will be extended to understand the basic ions chemistry that underpins these collisionally activated dissociation spectra. In parallel, we will use routine triple quadrupole mass spectrometry coupled via electrospray ionization with HLPLC. The low-energy collisionally induced fragmentations that are endemic to the triple quadrupole will be evaluated and compared with those occurring under the high-energy conditions of the four-sector instrument. The third approach is matrix-assisted laser desorption ionization (MALDI), which will be developed as a screening approach and as a structure-verification method by means of both molecular-ion and post- source=decomposition monitoring. In addition. MALDI capabilities will be extended to studies in which sites of modification and depurination in oligonucleotides must be located. The overall goal is to establish the merits of various mass spectrometric approaches to structure determination of DNA adducts, to understand the underlying ion chemistry, and to apply appropriately the methods to samples isolated from in vitro and in vivo studies.

最近我们已经确定高能串联质谱法是 对于确定 DNA 加合物的结构具有足够的特异性。 什么时候 然而，与快原子轰击质谱联用 检测限和灵敏度不足以支持调查 飞摩尔范围内的加合物。 为了克服这个限制，我们建议 开发和比较三种质谱方法 DNA加合物的灵敏度检测和结构测定。 串联 最近，具有扩展阵列检测功能的四扇区质谱法 在我们的实验室与电喷雾电离结合。 持续 开发是重中之重，因为产物离子谱已 足够的特异性来区分异构加合物。 努力将会 扩展以了解支撑这些的基本离子化学 碰撞激活解离谱。 同时，我们将使用 通过电喷雾耦合的常规三重四极杆质谱 用 HLPLC 进行电离。 低能碰撞引起的碎片 将评估和比较三重四极杆特有的 以及在四扇区的高能量条件下发生的情况 乐器。 第三种方法是基质辅助激光解吸 电离（MALDI），将被开发为一种筛选方法和 一种通过分子离子和后处理的结构验证方法 来源=分解监测。 此外。 MALDI 能力将 扩展到修饰和脱嘌呤位点的研究 必须定位寡核苷酸。 总体目标是建立 各种质谱结构测定方法的优点 DNA 加合物，了解基本的离子化学，并应用 适当地从体外和体内分离样品的方法 研究。