guanosine-monophosphate and 2--deoxyguanosine-3--phosphate

Many transition-metal complexes mediate DNA oxidation in the presence of oxidizing radiation, photosensitizers, or oxidants. The DNA oxidation products depend on the nature of the metal complex and the structure of the DNA. Earlier we reported trans-d,l-1,2-diaminocyclohexanetetrachloroplatinum (trans-Pt(d,l)(1,2-(NH(2))(2)C(6)H(10))Cl(4), [Pt(IV)Cl(4)(dach)]; dach = diaminocyclohexane) oxidizes 2'-deoxyguanosine 5'-monophosphate (5'-dGMP) to 7,8-dihydro-8-oxo-2'-deoxyguanosine 5'-monophosphate (8-oxo-5'-dGMP) stoichiometrically. In this paper we report that [Pt(IV)Cl(4)(dach)] also oxidizes 2'-deoxyguanosine 3'-monophosphate (3'-dGMP) stoichiometrically. The final oxidation product is not 8-oxo-3'-dGMP, but cyclic (5'-O-C8)-3'-dGMP. The reaction was studied by high-performance liquid chromatography, (1)H and (31)P nuclear magnetic resonance, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The proposed mechanism involves Pt(IV) binding to N7 of 3'-dGMP followed by nucleophilic attack of a 5'-hydroxyl oxygen to C8 of G and an inner-sphere, 2e(-) transfer to produce cyclic (5'-O-C8)-3'-dGMP and [Pt(II)Cl(2)(dach)]. The same mechanism applies to 5'-d[GTTTT]-3', where the 5'-dG is oxidized to cyclic (5'-O-C8)-dG. The Pt(IV) complex binds to N7 of guanine in cGMP, 9-Mxan, 5'-d[TTGTT]-3', and 5'-d[TTTTG]-3', but no subsequent transfer of electrons occurs in these. The results indicate that a good nucleophilic group at the 5' position is required for the redox reaction between guanosine and the Pt(IV) complex.

Topics: Cyclization; Deoxyguanine Nucleotides; Guanosine; Guanosine Monophosphate; Nuclear Magnetic Resonance, Biomolecular; Oligonucleotides; Organoplatinum Compounds; Oxidation-Reduction; Spectrophotometry, Ultraviolet; Spectroscopy, Fourier Transform Infrared

A 32P-postlabelling assay was developed for the analysis of adducts arising from the reaction of 2'-deoxyguanosine-3'-monophosphate with acetaldehyde, the primary oxidative metabolite of ethanol. The 32P-postlabelling reaction was optimized by testing various parameters such as the kinetics of phosphorylation by T4 polynucleotide kinase, substrate-concentration-dependent labelling efficiency and the concentration of the various ingredients of the phosphorylation reaction. The sensitivity to 3'-monophosphate dephosphorylation activity of nuclease P1 was also studied. Three stable adducts were separated by reversed-phase HPLC. The major stable adduct was structurally characterized and identified as N2-ethyl-2'-deoxyguanosine and could be detected, after reduction with NaBH4 or a mixture of ascorbic acid and GSH, in calf thymus DNA samples that had been reacted in vitro with acetaldehyde. DNA adducts were isolated after enzymatic digestion to mononucleotides followed by nuclease P1 digestion of normal nucleotides. The average levels of acetaldehyde-DNA adducts detected in these samples were 12.1 +/- 2.3 (n = 17) and 4.9 +/- 0.9 (n = 9) adducts/10(7) nucleotides after reduction with NaBH4, or ascorbic acid and GSH respectively. The 32P-postlabelling method was further validated by the detection of acetaldehyde adducts in liver DNA from mice treated with ethanol. The average concentration of the adducts detected in these animals was 1.5 +/- 0.8 (n = 7) adducts/10(8) nucleotides, as analyzed by reversed-phase HPLC with online detection of radioactivity.

Topics: Acetaldehyde; Animals; Cattle; Deoxyguanine Nucleotides; DNA; DNA Adducts; Ethanol; Guanosine Monophosphate; Isotope Labeling; Male; Mice; Mice, Inbred C57BL; Phosphorus Radioisotopes; Reference Standards; Spectrophotometry, Ultraviolet

Guanosine 3'-phosphate and 2'-deoxyguanosine 3'-phosphate are potent inhibitors of guanylate cyclase of various tissues of the silkworm, Bombyx mori. The inhibitory activity is specific for guanine 3'-nucleotides. Thus, 2'-GMP and 5'-GMP are one order or more less potent than 3'-GMP, and the other 3'-nucleotides except 3'-IMP, i.e. 3'-AMP, 2'-deoxy-3'-AMP, 3'-CMP, and 3'-TMP, have no inhibitory activity. Guanosine and 2'-deoxyguanosine are only weakly active. The 3'-nucleotide inhibition is dependent on the concentration of Mn2+. The concentrations of 2'-deoxy-3'-GMP and 3'-GMP required for 50% inhibition are 0.12 and 0.15 mM, respectively, with 5 mM Mn2+.

Topics: Adipose Tissue; Animals; Bombyx; Deoxyguanine Nucleotides; Guanine Nucleotides; Guanosine Monophosphate; Guanylate Cyclase; Kinetics; Male; Nucleosides; Nucleotides; Pupa; Structure-Activity Relationship