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 degreesC, 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-alpha Lys and dCyd-epsilon Lys, respectively). The yields at the reaction time of 72 h were 28.0% dCyd-alpha Lys, 13.4% dCyd-epsilon Lys, 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 degreesC for 24 h, 82% of the free dCyd-diazoate disappeared, indicating adduct formation with the polymer. At pH 7.4 and 37 degreesC, dCyd-alpha Lys and dCyd-ELys were fairly stable and gave rise to no product after incubation for 7 days. At pH 4.0 and 70 degreesC, 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 similar to1/3 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.