Nitric oxide-induced deamination of cytosine and guanine in deoxynucleosides and oligonucleotides.
The autoxidation of nitric oxide (NO.) forms the nitrosating agent N2O3, which can directly damage DNA by deamination of DNA bases following nitrosation of their primary amine functionalities. Within the G:C base pair, deamination results in the formation of xanthine and uracil, respectively. To determine the effect of DNA structure on the deamination of guanine and cytosine, the NO.-induced deamination rate constants for deoxynucleosides, single- and double-stranded oligonucleotides, and a G-quartet oligonucleotide were measured. Deamination rate constants were determined relative to morpholine using a Silastic membrane to deliver NO. at a rate of approximately 10-20 nmol/ml/min for 60 min, yielding a final concentration of approximately 600-1200 microM NO2-. GC/MS analysis revealed formation of nanomolar levels of deamination products from millimolar concentrations of deoxynucleosides and oligomers. Deamination rate constants for cytosine and guanine in all types of DNA were lower than the morpholine nitrosation rate constant by a factor of approximately 10(3)-10(4). Xanthine was formed at twice the rate of uracil, and this may have important consequences for mechanisms of NO.-induced mutations. Single-stranded oligomers were 5 times more reactive than deoxynucleosides toward N2O3. Double-stranded oligomers were 10-fold less reactive than single-stranded oligomers, suggesting that Watson-Crick base pairing protects DNA from deamination. G-quartet structures were also protective, presumably because of hydrogen bonding. These results demonstrate that DNA structure is an important factor in determining the reactivity of DNA bases with NO.-derived species.
Caulfield
J L
JL
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Wishnok
J S
JS
Tannenbaum
S R
SR
D003596:Cytosine / Q000737:chemistry*; D003641:Deamination; D006147:Guanine / Q000737:chemistry*; D006868:Hydrolysis; D007700:Kinetics; D009569:Nitric Oxide / Q000737:chemistry*; D009705:Nucleosides / Q000737:chemistry*; D009841:Oligonucleotides / Q000737:chemistry*
D009705:Nucleosides; D009841:Oligonucleotides; D009569:Nitric Oxide; D006147:Guanine; D003596:Cytosine