Masonic Cancer Center, University of Minnesota, 2231 6th Street SE, Minneapolis, Minnesota 55455, United States.
Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, United States.
Anal Chem. 2021 Apr 13;93(14):5754-5762. doi: 10.1021/acs.analchem.0c04895. Epub 2021 Apr 2.
Development of high-resolution/accurate mass liquid chromatography-coupled tandem mass spectrometry (LC-MS/MS) methodology enables the characterization of covalently modified DNA induced by interaction with genotoxic agents in complex biological samples. Constant neutral loss monitoring of 2'-deoxyribose or the nucleobases using data-dependent acquisition represents a powerful approach for the unbiased detection of DNA modifications (adducts). The lack of available bioinformatics tools necessitates manual processing of acquired spectral data and hampers high throughput application of these techniques. To address this limitation, we present an automated workflow for the detection and curation of putative DNA adducts by using diagnostic fragmentation filtering of LC-MS/MS experiments within the open-source software MZmine. The workflow utilizes a new feature detection algorithm, DFBuilder, which employs diagnostic fragmentation filtering using a user-defined list of fragmentation patterns to reproducibly generate feature lists for precursor ions of interest. The DFBuilder feature detection approach readily fits into a complete small-molecule discovery workflow and drastically reduces the processing time associated with analyzing DNA adductomics results. We validate our workflow using a mixture of authentic DNA adduct standards and demonstrate the effectiveness of our approach by reproducing and expanding the results of a previously published study of colibactin-induced DNA adducts. The reported workflow serves as a technique to assess the diagnostic potential of novel fragmentation pattern combinations for the unbiased detection of chemical classes of interest.
开发高分辨率/精确质量液相色谱-串联质谱 (LC-MS/MS) 方法可用于鉴定与遗传毒性剂相互作用后在复杂生物样品中诱导的共价修饰 DNA。使用数据依赖采集对 2'-脱氧核糖或核碱基进行恒定中性丢失监测是一种用于非选择性检测 DNA 修饰物（加合物）的强大方法。由于缺乏可用的生物信息学工具，需要对获得的光谱数据进行手动处理，从而阻碍了这些技术的高通量应用。为了解决这一限制，我们在开源软件 MZmine 中提出了一种自动化工作流程，用于通过在 LC-MS/MS 实验中使用诊断碎片过滤来检测和编纂可疑的 DNA 加合物。该工作流程利用了一种新的特征检测算法 DFBuilder，该算法使用用户定义的碎片模式列表进行诊断碎片过滤，可重现性地为感兴趣的前体离子生成特征列表。DFBuilder 特征检测方法很容易适应完整的小分子发现工作流程，并大大减少了分析 DNA 加合物组学结果相关的处理时间。我们使用真实的 DNA 加合物标准混合物验证了我们的工作流程，并通过重现和扩展先前发表的关于 colibactin 诱导的 DNA 加合物的研究结果证明了我们方法的有效性。所报告的工作流程可用于评估新型碎片模式组合用于非选择性检测感兴趣的化学类别的诊断潜力。
