Products of the reaction between a diazoate derivative of 2'-deoxycytidine and L-lysine and its implication for DNA-nucleoprotein cross-linking by NO or HNO(2).
Recently, we have reported that a stable diazoate intermediate (dCyd-diazoate) is produced upon the reaction of dCyd with nitrous acid and nitric oxide [Suzuki, T., Nakamura, T., Yamada, M., Ide, H., Kanaori, K., Tajima, K., Morii, T., and Makino, K. (1999) Biochemistry 38, 7151-7158]. In this work, the reaction of dCyd-diazoate with L-Lys was investigated. When 0.4 mM dCyd-diazoate was incubated with 10 mM L-Lys in sodium phosphate buffer (pH 7.4) at 37 degrees C, two unknown products were formed in addition to dUrd. By spectrometric measurements, the products were identified as dCyd-Lys adducts with C4(dCyd)-N(alpha)(Lys) and C4(dCyd)-N(epsilon)(Lys) linkages (abbreviated as dCyd-alphaLys and dCyd-epsilonLys, respectively). The yields at the reaction time of 72 h were 28.0% dCyd-alphaLys, 13.4% dCyd-epsilonLys, and 11.1% dUrd with 33.9% unreacted dCyd-diazoate. When 0.4 mM dCyd-diazoate was incubated with 22 mg/mL poly(L-Lys) at pH 7.4 and 37 degrees C for 24 h, 82% of the free dCyd-diazoate disappeared, indicating adduct formation with the polymer. At pH 7.4 and 37 degrees C, dCyd-alphaLys and dCyd-epsilonLys were fairly stable and gave rise to no product after incubation for 7 days. At pH 4.0 and 70 degrees C, both adducts disappeared with the same first-order rate constant of 1.7 x 10(-)(6) s(-)(1) (t(1/2) = 110 h), which was approximately (1)/(3) of that of dCyd. These results suggest that if dCyd-diazoate is formed in DNA in vivo, it may react with free L-Lys and the side chain of L-Lys in nucleoproteins, resulting in stable adducts and DNA-protein cross-links, respectively.
Suzuki
T
T
Institute of Advanced Energy, Kyoto University, Gokasho, Uji 611-0011, Japan.
Yamada
M
M
Nakamura
T
T
Ide
H
H
Kanaori
K
K
Tajima
K
K
Morii
T
T
Makino
K
K
D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents; D018736:DNA Adducts / Q000737:chemistry*; D003979:Diazonium Compounds / Q000737:chemistry*; D008239:Lysine / Q000737:chemistry*; D009569:Nitric Oxide / Q000737:chemistry*; D009608:Nitrous Acid / Q000737:chemistry*; D011743:Pyrimidines / Q000737:chemistry*
C119435:2'-deoxyribofuranosyl-2-oxopyrimidine-4-diazoate; D003432:Cross-Linking Reagents; D018736:DNA Adducts; D003979:Diazonium Compounds; D011743:Pyrimidines; D009569:Nitric Oxide; D008239:Lysine; D009608:Nitrous Acid