NMR Solution Structures of Clustered Abasic Site Lesions in DNA: Structural Differences between 3′-Staggered (−3) and 5′-Staggered (+3) Bistranded Lesions

摘要

Ionizing radiation produces a distinctive pattern of bistranded clustered lesions in DNA. Anrelatively low number of clustered lesions may be lethal to cells when compared to a larger number of singlenlesions. Enzyme cleavage experiments suggest that the orientation of bistranded lesions causes differentialnrecognition and removal of these lesions. Like that of a previous study of bistranded abasic site lesion [Hazel,nR.D., Tian, K., and de los Santos, C. (2008) Biochemistry 47, 11909-11919], the aimof this investigation wasnto determine the structures of two DNA duplexes each containing two synthetic apurinic/apyrimidinic (AP)nresidues, positioned on opposite strands and separated by two base pairs. In the first duplex, the AP residuesnare staggered in the 30norientation [-3 duplex, (AP)2-3 duplex], while in the second duplex, the AP residuesnare staggered in the 50norientation [þ3 duplex, (AP)2þ3 duplex]. NOESY spectra recorded in 100 and 10%nD2O buffer solutions allowed the assignment of the nonexchangeable and exchangeable protons, respectively,nfor each duplex. Cross-peak connectivity in the nonexchangeable proton spectra indicates that the duplex is anregular right-handed helix with the AP residues and orphan bases located inside the duplexes. Thenexchangeable proton spectra establish the formation of Watson-Crick G3nC alignment for the two basenpairs between the lesion sites in both duplexes.Distance-restrainedmolecular dynamics simulation confirmednthe intrahelical orientations of the AP residues. The proximity of the AP residues across the minor groovenof the -3 duplex and across the major groove in the þ3 duplex is similar to their locations in the case of -1nand þ1 clusters. This difference in structure may be a key factor in the differential recognition of bistrandednAP lesions by human AP endonuclease.