Base-Displaced Intercalated Structure of the 3-(2-Deoxy-β-D-erythropentofuranosyl)-pyrimido[1,2- f ]purine-6,10(3 H ,5 H )-dione (6-oxo-M 1 dG) Lesion in DNA.

The genotoxic 3-(2-deoxy-β-D-erythro-pentofuranosyl)pyrimido[1,2-α]purin-10(3H)-one (M 1 dG) DNA lesion arises from endogenous exposures to base propenals generated by oxidative damage and from exposures to malondialdehyde (MDA), produced by lipid peroxidation. Once formed, M 1 dG may oxidize, in vivo , to 3-(2-deoxy-β-D-erythropentofuranosyl)-pyrimido[1,2- f ]purine-6,10(3 H ,5 H )-dione (6-oxo-M 1 dG). The latter blocks DNA replication and is a substrate for error-prone mutagenic bypass by the Y-family DNA polymerase hpol η. To examine structural consequences of 6-oxo-M 1 dG damage in DNA, we conducted NMR studies of 6-oxo-M 1 dG incorporated site-specifically into 5' -d(C 1 A 2 T 3 X 4 A 5 T 6 G 7 A 8 C 9 G 10 C 11 T 12 )-3':5'-d(A 13 G 14 C 15 G 16 T 17 C 18 A 19 T 20 C 21 A 22 T 23 G 24 )-3' ( X = 6-oxo-M 1 dG). NMR spectra afforded detailed resonance assignments. Chemical shift analyses revealed that nucleobase C 21 , complementary to 6-oxo-M 1 dG, was deshielded compared with the unmodified duplex. Sequential NOEs between 6-oxo-M 1 dG and A 5 were disrupted, as well as NOEs between T 20 and C 21 in the complementary strand. The structure of the 6-oxo-M 1 dG modified DNA duplex was refined by using molecular dynamics (rMD) calculations restrained by NOE data. It revealed that 6-oxo-M 1 dG intercalated into the duplex and remained in the anti -conformation about the glycosyl bond. The complementary cytosine C 21 extruded into the major groove, accommodating the intercalated 6-oxo-M 1 dG. The 6-oxo-M 1 dG H7 and H8 protons faced toward the major groove, while the 6-oxo-M 1 dG imidazole proton H2 faced into the major groove. Structural perturbations to dsDNA were limited to the 6-oxo-M 1 dG damaged base pair and the flanking T 3 :A 22 and A 5 :T 20 base pairs. Both neighboring base pairs remained within the Watson-Crick hydrogen bonding contact. The 6-oxo-M 1 dG did not stack well with the 5'-neighboring base pair T 3 :A 22 but showed improved stacking with the 3'-neighboring base pair A 5 :T 20 . Overall, the base-displaced intercalated structure was consistent with thermal destabilization of the 6-oxo-M 1 dG damaged DNA duplex; thermal melting temperature data showed a 15 °C decrease in T m compared to the unmodified duplex. The structural consequences of 6-oxo-M 1 dG formation in DNA are evaluated in the context of the chemical biology of this lesion.