1. Electrons and DNA: From Electron Attachment to DNA Strand Breaks Michael Sevilla Chemistry Department Oakland University

2. DNA electron and hole scavenging G +, CH, T –, Sug electron and hole scavenger: electron and hole scavenger: S G+, electron scavenger G+, S, electron scavenger CH, T–, hole scavenger CH, T–, hole scavenger

3. What are the mechanisms of electrons reaction with DNA?

4. DNA and Electrons A variety of studies show that aqueous electrons add to the DNA bases and do not cause strand breaks. The base anion radicals undergo protonation reactions. Only nonsolvated electrons with kinetic energy (LEE) cause DNA strand breaks. At low temperatures electron in DNA are stable for long times.

5.  (Å - 1 ) 0.75 0.92 1.4 2.8 Vacuum pdIdCpdIdC DNA (salmon sperm) pdAdTpdAdT Electron Tunneling  Decay Constant in DNA at 77 K k = 10 11 exp(-  D) Z Cai and M Sevilla, in "Topics in Current Chemistry 237: Long Range Transfer in DNA II", Gary Shuster, Ed., Springer-Verlag, pages 103-128, 2004.

6.  DNA  irradiation Studies  Irradiate hydrated DNA at 77K in frozen solutions Direct Ionization of DNA results Identify initial radicals by Electron Spin Resonance Spectroscopy Ionizing radiation DNA + + DNA – DNA Oxidative pathReductive path "The Chemical Consequences of Radiation Damage to DNA" D. Becker and M. D. Sevilla, in Advances in Radiation Biology, Vol.17, (J. Lett, Ed.) Academic Press, 121-180 (1993).

7. RADICAL COMPOSITION (  - irradiated DNA ) ESR Spectra The composition of DNA radicals (77 K) is ca.: 25% (C-) C(N3)H 25% T- 40% G+ 10% Sugar All to ±5% C(N3)H

8. Effect of scavengers on G-values L. I. Shukla, R. Pazdro, D. Becker and M. D. Sevilla, Radiation Research: Vol. 163, 59-602 (2005). Sug

9. Hole induced Sugar radicals in DNA Oxidative Pathway C1’ C3’ C4’ ? C5’

10. Oxidation pathway: formation of neutral sugar radical(s) from sugar cation radical(s) Deprotonation from C1’, (C2’), C3’, C4’, C5’ may occur. L. I. Shukla, R Pazdro, D Becker and M D Sevilla, RAD. RES.163, 591–602 (2005)

11. How do electrons damage DNA? Clues from Ion Beam Irradiation of DNA ca. 100 MeV/nucleon O +8, Ar +18, Kr +36, Xe +54 D Becker, A Bryant-Friedrich, C Trzasko and M D Sevilla, Rad Res 160, 174 (2003)

12. PARTRAC Calculations, Hauptner et al (2006)

13. Cross Section of Heavy Ion Track Aloke Chatterjee and John Magee, J. Phys. Chem. Vol. 84, 3629-3536 (1980)

14. High Yields of Sugar radicals found in ion-beam irradiated DNA D. Becker, A. Bryant-Friedrich, C. Trzasko, and M. D. Sevilla, Radiation Research, 160, 174-185 (2003) M. Bowman, D.Becker, M. D. Sevilla and J. Zimbrick, Radiation Research: Vol. 163, 447-454 (2005). Excited States Important?

15. Yields of Sugar Radicals Increased in core of ion beam irradiated DNA ● Sugar Radical Yield per Joule in track core more than in the penumbra. ● New sugar phosphate species found in ion beam irradiated DNA D. Becker, A. Bryant-Friedrich, C. Trzasko, and M. D. Sevilla, Radiation Research, 160, 174-185 (2003)

16. Ar ion Irradiated DNA Phosphoryl radicals - strand break radical(s)

17. 2 nd Strand Break radical found in ion beam irradiated DNA N from DNA Simulation using theoretical spectral parameters C3’ dephos ●

18. LEE Induced Strand Scission Path B bond scission Path A bond scission LEE D. Becker, A. Bryant-Friedrich, C. Trzasko, and M. D. Sevilla, Radiation Research, 160, 174-185 (2003)

19. What about Excited states? We know excitation of DNA holes forms sugar radicals

20. Visible excitation converts G + to Sugar Radicals A Adhikary, S Collins, D Khanduri, and M D Sevilla, JPC B 2007, 111, 7415-7421 A Adhikary, A Kumar and M D Sevilla, Rad Res 2006 165, 479–484

21. LEE Reaction with DNA: Strand Break Formation LEE capture at the base or the sugar phosphate backbone? What energies are needed? Are excited states involved?

22. B. Boudaiffa, P. Cloutier, D. Hunting, M. A. Huels, L. Sanche, Science 2000, 287, 1658 F. Martin, P.D. Burrow, Z. Cai, P. Cloutier, D.J. Hunting, L. Sanche, Phys. Rev. Lett. 93, 068101-1 (2004) DNA Strand break induced by low energy electrons

23. DNA Modeling the LEE Induced DNA Strand Break Li, X.; Sevilla, M. D.; Sanche, L.; J. Am. Chem. Soc., 2003, 125, 13668 Sugar-Phosphate-Sugar model

24. The Potential Energy Surfaces DNA

25. Basis set dependence of Spin Distribution 3-21G(d) 6-31G(d) 6-31+G(d) Xifeng Li and Michael D. Sevilla, in Theoretical Treatment of the Interaction of Radiation with Biological Systems. J. R. Sabin and E. Brandas, Eds., Advances in Quantum Chemistry, Elsevier, Volume 52, 59-88 (2007).

26. New DFT Calculations on 5’-dTMPH Anion Radical What about the vertical surface? A. Kumar; M. Sevilla, J. Phys. Chem. B 2007, 111, 5464-5474 Recent Previous Work : J. Simons, Acc. Chem. Res. 2006, 39, 772-779 Bao, X.; Wang, J.; Gu, J.; Leszczynski, J. Proc. Nat. Acad. Sci.U.S.A. 2006, 103, 5658. Gu, J.; Wang, J.; Leszczynski, J. J. Am. Chem. Soc. 2006, 128, 9322. Adiabatic Barriers of 7 kcal/mol for 3’- C-O bond and 14 kcal/mol for 5’-C-O bond

27. LEE Induced strand break in 5'-dTMPH as a model: Vertical or Adiabatic pathways? vertical adiabatic

28. B3LYP/6-31G* calculated adiabatic and vertical potential surfaces (PES) of C5'-O5' bond dissociation of 5'-dTMPH radical anion. The singly occupied molecular orbital (SOMO) is also shown 1.45 1.5 1.6 1.7 1.8 1.9 2.0 B3LYP/6-31G * kcal/mol 26.0 24.0 22.0 20.0 18.0 16.0 14.0 12.0 10.0 8.0 6.0 4.0 2.0 0.0 -8.0 C 5' -O 5' (Å) Vertical PES 16.5 17.6 22.5 25.5 24.4 20.7 15.5 0.7 4.8 10.8 14.8 1.78 Å -7.4 TS Adiabatic PES

29. B3LYP/6-31++G** calculated adiabatic and vertical potential energy surfaces (PES) of C5'-O5' bond dissociation of 5'-dTMPH radical anion. B3LYP/6-31++G ** kcal/mol 4.8 5.7 10.9 18.0 21.7 19.1 14.4 0.6 4.5 10.2 13.5 1.78 Å 0.5 -5.8 20.0 18.0 16.0 14.0 12.0 10.0 8.0 6.0 4.0 2.0 0.0 -2.0 -4.0 -6.0 8.2 kcal/mol 22.0 TS Adiabatic PES Vertical PES 1.45 1.5 1.6 1.7 1.8 1.9 2.0 C 5' -O 5' (Å)

30. B3LYP/6-31G** optimized geometries of neutral and anionic radical of 5'-dTMP with Na+ and 11 H2O. A. Kumar; M. Sevilla, J. Phys. Chem. B 2007, 111, 5464-5474 5'-dTMPNa + 11 H2O (Neutral)5'-dTMPNa + 11 H2O (Anion radical) Na+

31. DFT (B3LYP/6-31G**) calculated adiabatic potential energy surface (PES) of C5'-O5' bond dissociation of hydrated (11 H2O) 5'-dTMP radical anion with Na+ as a counter ion. Na + B3LYP/6-31G ** kcal/mol 33.0 30.0 27.0 24.0 21.0 18.0 15.0 12.0 9.0 6.0 3.0 0.0 -6.0 1.5 1.6 1.7 1.8 1.9 2.0 2.1 2.2 2.3 2.5 C 5' -O 5' (Å) 15.4 28.9 11.0 -3.3 Na +

32. Proposed mechanism of single strand break (SSB) due to attachment of LEE with 5'-dTMPH molecule

33. Why C-O not P-O? Experimentally only cleavage at the C-O bond is found. Y Zheng, P Cloutier, D Hunting, J R Wagner, L Sanche, JCP, 124, 064710 (2006). The loss of phosphate anion is the driving force for the reaction because the phosphate radical has a such a large EA. J. Simons, Acc. Chem. Res. 2006, 39, 772-779

34. Base Release in Nucleosides Induced by Low Energy Electrons X. Li, L. Sanche and M. Sevilla, Radiation Research, 165, 721 (2006) J. Gu, Y. Xie and H. F. Schaefer, III, J. Am. Chem. Soc. 127, 1053– 1057 (2005).

35. Potential Energy Surfaces (PESs) for C-N bond dissociation in Nucleoside Anion Radicals: Calculated with DFT (b3lyp, 6-31+G(d)) dA and dG anion radicals dT and dC Anion Radicals X. Li, L. Sanche and M. Sevilla, Radiation Research, 165, 721 (2006)

36. LEE Induced Excited States LEE induced resonances found experimentally suggest excited state involvement. What are the available excited state levels? We performed TD-DFT calculations to aid our understanding in 5’-dTMP - ●

37. J. Simons, Acc. Chem. Res. 2006, 39, 772-779 J. Berdys, I. Anusiewicz, P. Skurski, J. Simons JACS (2004) Excited States of Sugar Phosphate Portion

38. Molecular orbital Energies and MO plots of neutral 5'-dTMPH. B3LYP/6- 31G*. Scaled values by method of Modelli and Jones JPC 2006. *Experimental VOEs of thymine (Aflatooni et al. J. Phys. Chem. A (1998) 102, 6205 ) MOs Orbital Energy (eV) Scaled VOE (eV) LUMO + 4 (σ 3 *) LUMO + 3 (σ 2 *) LUMO + 2 (σ 1 *) LUMO + 1 (π 2 *) LUMO (π 1 *) HOMO (π) 1.78 2.64 1.27 2.23 0.73 1.80 0.43 1.56 (1.71)* -0.84 0.53 (0.29)* -6.24 ca. - 5 eV

39. Summary Strand break formation by LEE induced C-O bond scission is the lowest energy pathway in comparison to C-N, N-H or C-H bond scissions. LEE induced anion excited states likely provide a facile route to strand breaks.

40. Ph.D. Students. Amitava Adhikary Deepti Khanduri Anil Kumar Sean Collins David Becker Alyson Engle Lata Shukla Tom Casey Collaborators Leon Sanche Xifeng Li Acknowledgments This research is supported by NIH NCI RO1CA045424 DNA Radiation Chemistry Group at Oakland University

41. OU ESR GROUP