Comprehensive analysis of DNA modifications using data-independent SIM/MS2 acquisition and wSIM-City

A novel software has been created to comprehensively characterize covalent modifications of DNA through mass spectral analysis of enzymatically hydrolyzed DNA using the neutral loss of 2′-deoxyribose, a nearly universal MS2 fragmentation process of protonated 2′-deoxyribonucleosides. These covalent modifications termed DNA adducts form through xenobiotic exposures or by reaction with endogenous electrophiles and can induce mutations during cell division and initiate carcinogenesis. DNA adducts are typically present at trace levels in the human genome, requiring a very sensitive and comprehensive data acquisition and analysis method. Our software, wSIM-City, was created to process mass spectral data acquired by wide selected ion monitoring (wSIM) with gas-phase fractionation (GPF) and coupled to wide MS2 fragmentation. This untargeted approach can detect DNA adducts at trace levels as low as 1.5 adducts per 109 nucleotides. This level of sensitivity is sufficient for comprehensive analysis and characterization of DNA modifications in human specimens.本研究开发了一款新型软件，可通过对酶解DNA进行质谱分析，依托质子化2'-脱氧核苷（protonated 2′-deoxyribonucleosides）近乎通用的MS2碎裂过程——2'-脱氧核糖中性丢失（neutral loss of 2′-deoxyribose），全面表征DNA的共价修饰。这类被称为DNA加合物（DNA adducts）的共价修饰可通过外源性暴露或与内源性亲电试剂反应形成，能够在细胞分裂过程中诱发突变并启动致癌进程。DNA加合物在人类基因组中通常以痕量水平存在，因此需要具备极高灵敏度的全面数据采集与分析方法。本软件wSIM-City专为处理经宽范围选择离子监测（wide selected ion monitoring, wSIM）结合气相分级分离（gas-phase fractionation, GPF）并耦合宽范围MS2碎裂所采集的质谱数据而开发。该非靶向分析方法可检测到低至每10^9个核苷酸1.5个加合物的痕量水平DNA加合物，这一灵敏度足以满足人类样本中DNA修饰的全面分析与表征需求。