Accurate and rapid modeling of iron-bleomycin-induced DNA damage using tethered duplex oligonucleotides and electrospray ionization ion trap mass spectrometric analysis.
Accurate and rapid modeling of iron-bleomycin-induced DNA damage using tethered duplex oligonucleotides and electrospray ionization ion trap mass spectrometric analysis.
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使用系链双链寡核苷酸和电喷雾电离离子阱质谱分析对铁博来霉素诱导的 DNA 损伤进行准确、快速的建模。
DOI：
10.1093/nar/28.9.1978
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发表时间：
2000
影响因子：
14.9
通讯作者：
Vouros,P
中科院分区：
文献类型：
--
作者：
Harsch,A;Marzilli,LA;Bunt,RC;Stubbe,J;Vouros,P
Bleomycin B2(BLM) in the presence of iron [Fe(II)] and O2catalyzes single-stranded (ss) and double-stranded (ds) cleavage of DNA. Electrospray ionization ion trap mass spectrometry was used to monitor these cleavage processes. Two duplex oligonucleotides containing an ethylene oxide tether between both strands were used in this investigation, allowing facile monitoring of all ss and ds cleavage events. A sequence for site-specific binding and cleavage by Fe–BLM was incorporated into each analyte. One of these core sequences, GTAC, is a known hot-spot for ds cleavage, while the other sequence, GGCC, is a hot-spot for ss cleavage. Incubation of each oligonucleotide under anerobic conditions with Fe(II)–BLM allowed detection of the non-covalent ternary Fe–BLM/oligonucleotide complex in the gas phase. Cleavage studies were then performed utilizing O2-activated Fe(II)–BLM. No work-up or separation steps were required and direct MS and MS/MS analyses of the crude reaction mixtures confirmed sequence-specific Fe–BLM-induced cleavage. Comparison of the cleavage patterns for both oligonucleotides revealed sequence-dependent preferences for ss and ds cleavages in accordance with previously established gel electrophoresis analysis of hairpin oligonucleotides. This novel methodology allowed direct, rapid and accurate determination of cleavage profiles of model duplex oligonucleotides after exposure to activated Fe–BLM.
