A new mass spectrometric approach to detect modifications in DNA.
A new approach is described for the enzymatic digestion of DNA yielding oligonucleotides ranging from dinucleoside monophosphates to octanucleoside heptaphosphates. DNA was digested by means of the benzon nuclease, an unspecific nuclease, and alkaline phosphatase to remove the terminal phosphate. The mixture of oligonucleotides was separated using capillary-zone electrophoresis with a buffer system, yielding a rather strong electro-osmotic flow. The oligomers are separated into groups with nucleotides of the same chain length. The separation capillary was used as the innermost capillary of an electrospray spraying system. Negative molecular ions of the nucleotides were recorded using a home-built interface and ion source for a sector-field mass spectrometer. This approach allows the facile detection of DNA modifications since they lead not only to differences in mass, but also can possess altered electrophoretic mobility. For modifying reactions which exhibit sequence specificity, the information is retained in the oligomers. Thus, reactions of DNA with electrophiles can be evaluated at different levels, since in longer chains, even complex sequence specificity may be apparent. Results from calf thymus DNA digests and preliminary experiments with DNA adducts with styrene oxide are discussed.
Janning
P
P
ISAS Institut für Spektrochemie und Angewandte Spektroskopie, Dortmund, Germany.
Schrader
W
W
Linscheid
M
M
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000032:analysis* / Q000302:isolation & purification; D018736:DNA Adducts / Q000032:analysis; D004586:Electrophoresis; D004852:Epoxy Compounds; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000032:analysis / Q000302:isolation & purification; D013056:Spectrophotometry, Ultraviolet
D018736:DNA Adducts; D004852:Epoxy Compounds; D009841:Oligonucleotides; D004247:DNA; C013690:styrene oxide