Book of Abstracts: Albany 2003
June 17-21 2003
NMR Solution Structure of Bistranded Furan Abasic Site Lesions: Orientation Matters
Ionizing radiation has the distinction in that it can produce multiple damage sites (MDS) in DNA, which can be clustered within 1 ? 2 turns of the DNA helix. A relatively low number of such clustered lesions may prove to be lethal as compared to a larger number of single lesions. Enzyme kinetics experiments done in our lab and elsewhere suggest that the orientation of clustered lesions influences the efficiency with which such lesions can be repaired. We have solved the structures of two pairs of DNA Duplexes each of which contains clustered abasic site damage represented by a pair of furan lesions.
In the first case, we determined the structures of a pair of 13-mer duplexes containing a pair of furan residues positioned on opposite strands and staggered 1 base pair 3? or 5? to the furan residue on the complimentary strand. NOESY spectra were collected for both duplexes in D2O and H2O. This allowed for the assignment of non-exchangeable and exchangeable protons. For both the ?1 and +1 clustered lesions, cross peak connectivity in the spectra of non-exchangeable protons indicated that the helix collapsed at the lesion site. The spectra for exchangeable protons indicated the formation of a Watson-Crick G-C base pair at the lesion site in both duplexes. Distance restrained molecular dynamics simulation was used to determine the solution structures of all duplexes. Sets of 20 structures were produced for each duplex, and those structures that exhibited a pair wise rmsd of 0.75 ? 1.15 A were chosen as representative final structures. These structures showed that furan residues were extruded towards the minor groove in the case of the ?1 clustered arrangement but the furans were extruded towards the major groove in the +1 cluster. The three-dimensional structures can be correlated with the incision activity of AP endonucleases, which remove such lesions during DNA repair.
Key words: DNA damage, multiple damage site, furan, clustered lesion, ionizing radiation.
Raphael D. Hazel1
1Department of Physiology & Biophysics