No abstract

A functional human homologue of Escherichia coli endonuclease III (Nth-Eco protein) has recently been cloned and characterized [Aspinwall, R., Rothwell, D. G., Roldan-Arjona, T., Anselmino, C., Ward, C. J., Cheadle, J. P., Sampson, J. R., Lindahl, T., Harris, P. C., and Hickson, I. D. (1997) Proc. Natl. Acad. Sci. U.S.A., 94, 109-114]. This enzyme, designated hNTH1 protein, shares an extensive sequence similarity with Nth-Eco protein and a related enzyme from Schizosaccharomyces pombe (Nth-Spo protein). We investigated the substrate specificity of this human enzyme for oxidative DNA base damage, using the technique of gas chromatography/isotope-dilution mass spectrometry. Four different DNA substrates damaged by various free radical-generating systems were used. 5-Hydroxycytosine, thymine glycol, 5-hydroxy-6-hydrothymine, 5,6-dihydroxycytosine, and 5-hydroxyuracil were substrates of hNTH1 protein among 17 lesions found in DNA substrates. The substrate specificity and excision kinetics of the human enzyme were found to be significantly different from those of Nth-Spo and Nth-Eco proteins.

A flash photolysis-resonance fluorescence technique was used to study the rate constant for the reaction of OH radicals with dimethyl carbonate over the temperature range 252-370 K. The rate constant exhibited a weak temperature dependence, increasing at both low and high temperature from a minimum value of approximately 3.1 × 10 -13 cm 3 molecule -1 s -1 near room temperature. Pulse radiolysis/transient UV absorption techniques were used to study the ultraviolet absorption spectra and kinetics of CH 3 OC(O)OCH 2 and CH 3 OC(O)CH 2 O 2 radicals at 296 K. Absorption cross sections of CH 3 OC(O)OCH 2 and CH 3 OC(O)OCH 2 O 2 at 250 nm were (3.16 ( 0.34) × 10 -18 and (3.04 ( 0.43) × 10 -18 cm 2 molecule -1 , respectively. Rate constants measured for the self-reactions of CH 3 OC(O)OCH 2 and CH 3 OC(O)OCH 2 O 2 radicals and reactions of CH 3 OC(O)OCH 2 O 2 radicals with NO and NO 2 were (5.6 ( 1.1) × 10 -11 , (1.27 ( 0.21) × 10 -11 , (1.2 ( 0.2) × 10 -11 , and (1.2 ( 0.2) × 10 -11 cm 3 molecule -1 s -1 , respectively. The rate constant for reaction of F atoms with dimethyl carbonate was determined by a pulse radiolysis absolute rate technique to be (6.1 ( 0.9) × 10 -11 cm 3 molecule -1 s -1 . A FTIR smog chamber system was used to show that, in 760 Torr of air at 296 K, CH 3 OC(O)OCH 2 O radicals are lost via three competing processes: 42 ( 15% via reaction with O 2 , 14 ( 2% via H atom elimination, and 44 ( 10% via decomposition and/or isomerization. Relative rate techniques were used to measure rate constants for the reactions of F atoms with CH 3 OC(O)OCH 3 , (6.4 ( 1.4) × 10 -11 cm 3 molecule -1 s -1 , and Cl atoms with CH 3 OC(O)OCH 3 , CH 3 OC(O)OCH 2 Cl, CH 3 OC(O)-OCHO, and HC(O)OC(O)OCHO, (2.3 ( 0.8) × 10 -12 , (4.6 ( 2.8) × 10 -13 , (1.7 ( 0.1) × 10 -13 , and (1.7 ( 0.1) × 10 -14 cm 3 molecule -1 s -1 , respectively. Results are discussed in the context of the atmospheric chemistry of CH 3 OC(O)OCH 3 .