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

The genotoxic 3-(2-deoxy-β-D-erythro-pentofuranosyl)pyrimido[1,2-α]purin-10(3H)-one (MdG) 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, MdG may oxidize, , to 3-(2-deoxy-β-D-erythropentofuranosyl)-pyrimido[1,2-]purine-6,10(3,5)-dione (6-oxo-MdG). 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-MdG damage in DNA, we conducted NMR studies of 6-oxo-MdG incorporated site-specifically into 5' -d(CATATGACGCT)-3':5'-d(AGCGTCATCATG)-3' ( = 6-oxo-MdG). NMR spectra afforded detailed resonance assignments. Chemical shift analyses revealed that nucleobase C, complementary to 6-oxo-MdG, was deshielded compared with the unmodified duplex. Sequential NOEs between 6-oxo-MdG and A were disrupted, as well as NOEs between T and C in the complementary strand. The structure of the 6-oxo-MdG modified DNA duplex was refined by using molecular dynamics (rMD) calculations restrained by NOE data. It revealed that 6-oxo-MdG intercalated into the duplex and remained in the -conformation about the glycosyl bond. The complementary cytosine C extruded into the major groove, accommodating the intercalated 6-oxo-MdG. The 6-oxo-MdG H7 and H8 protons faced toward the major groove, while the 6-oxo-MdG imidazole proton H2 faced into the major groove. Structural perturbations to dsDNA were limited to the 6-oxo-MdG damaged base pair and the flanking T:A and A:T base pairs. Both neighboring base pairs remained within the Watson-Crick hydrogen bonding contact. The 6-oxo-MdG did not stack well with the 5'-neighboring base pair T:A but showed improved stacking with the 3'-neighboring base pair A:T. Overall, the base-displaced intercalated structure was consistent with thermal destabilization of the 6-oxo-MdG damaged DNA duplex; thermal melting temperature data showed a 15 °C decrease in compared to the unmodified duplex. The structural consequences of 6-oxo-MdG formation in DNA are evaluated in the context of the chemical biology of this lesion.