NDLI: Accurate and rapid modeling of iron–bleomycin-induced DNA damage using tethered duplex oligonucleotides and electrospray ionization ion trap mass spectrometric analysis

Content Provider	PubMed Central
Author	Harsch, Andreas Marzilli, Lisa A. Bunt, Richard C. Stubbe, Joanne Vouros, Paul
Copyright Year	2000
Abstract	Bleomycin B2 (BLM) in the presence of iron [Fe(II)] and O2 catalyzes 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.
Starting Page	1978
File Format	PDF
ISSN	13624962
e-ISSN	13624962
Journal	Nucleic Acids Research
Issue Number	9
Volume Number	28
Language	English
Publisher	Oxford University Press
Publisher Date	2000-05-01
Access Restriction	Open
Rights Holder	Oxford University Press
Subject Keyword	Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Genetics

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