JBIC Journal of Biological Inorganic Chemistry

, Volume 15, Issue 2, pp 175–182 | Cite as

ENDOR and ESEEM investigation of the Ni-containing superoxide dismutase

Original Paper


Superoxide dismutases (SODs) protect cells against oxidative stress by disproportionating O2 to H2O2 and O2. The recent finding of a nickel-containing SOD (Ni-SOD) has widened the diversity of SODs in terms of metal contents and SOD catalytic mechanisms. The coordination and geometrical structure of the metal site and the related electronic structure are the keys to understanding the dismutase mechanism of the enzyme. We performed Q-band 14N,1/2H continuous wave (CW) and pulsed electron–nuclear double resonance (ENDOR) and X-band 14N electron spin echo envelope modulation (ESEEM) on the resting-state Ni-SOD extracted from Streptomyces seoulensis. In-depth analysis of the data obtained from the multifrequency advanced electron paramagnetic resonance techniques detailed the electronic structure of the active site of Ni-SOD. The analysis of the field-dependent Q-band 14N CW ENDOR yielded the nuclear hyperfine and quadrupole coupling tensors of the axial Nδ of the His-1 imidazole ligand. The tensors are coaxial with the g-tensor frame, implying the g-tensor direction is modulated by the imidazole plane. X-band 14N ESEEM characterized the hyperfine coupling of Nε of His-1 imidazole. The nuclear quadrupole coupling constant of the nitrogen suggests that the hydrogen-bonding between Nε–H and OGlu-17 present for the reduced-state Ni-SOD is weakened or broken upon oxidizing the enzyme. Q-band 1H CW ENDOR and pulsed 2H Mims ENDOR showed a strong hyperfine coupling to the protons(s) of the equatorially coordinated His-1 amine and a weak hyperfine coupling to either the proton(s) of a water in the pocket at the side opposite the axial Nδ or the proton of a water hydrogen-bonded to the equatorial thiolate ligand.


Nickel-containing superoxide dismutase Electron paramagnetic resonance Electron-nuclear double resonance Electron spin echo envelope modulation 



Copper- and zinc-containing superoxide dismutase


Continuous wave


Electron–nuclear double resonance


Electron paramagnetic resonance


Electron spin echo envelope modulation


Nickel-containing superoxide dismutase


Superoxide dismutase



This work was supported by the Korea Research Foundation (KRF-2006-312-C00219, H.I.L.), the Research Fellowship of BK21 project (S.O.K.), and the NSF (MCB0723330, B.M.H.).


  1. 1.
    Fridovich I (1997) J Biol Chem 272:18515–18517CrossRefPubMedGoogle Scholar
  2. 2.
    Cabelli DE, Riley D, Rodriguez JA, Valentine JS, Zhu H (2000). In: Meunier B (ed) Biomimetic oxidations catalyzed by transition metal complexes. Imperial College Press, London, chap 10 Google Scholar
  3. 3.
    Miller A-F (2003) In: Que L Jr, Tolman W (eds) Coordination chemistry in the biosphere and geosphere. Pergamon, Amsterdam, pp 479–506Google Scholar
  4. 4.
    Miller A-F (2004) Curr Opin Chem Biol 8:162–168CrossRefPubMedGoogle Scholar
  5. 5.
    Tainer JA, Getzoff ED, Richardson JS, Richardson DC (1983) Nature 306:284–287CrossRefPubMedGoogle Scholar
  6. 6.
    Bordo D, Djinovic-Carugo K, Bolognesi M (1994) J Mol Biol 238:366–368CrossRefPubMedGoogle Scholar
  7. 7.
    Djinovic-Carugo K, Battistoni A, Carri M, Polticelli F, Desideri A, Rotilio G, Coda A, Wilson K, Bolognesi M (1996) Acta Crystallogr D 52:176–188CrossRefPubMedGoogle Scholar
  8. 8.
    Bordo D, Matak D, Djinovic-Carugo K, Rosano C, Pesce A, Bolognesi M, Stoppolo ME, Falconi M, Battistoni A, Desideri A (1999) J Mol Biol 285:283–296CrossRefPubMedGoogle Scholar
  9. 9.
    Parker MW, Blake CC (1988) J Mol Biol 199:649–661CrossRefPubMedGoogle Scholar
  10. 10.
    Ludwig ML, Metzger AL, Pattridge KA, Stallings WC (1991) J Mol Biol 219:335–358CrossRefPubMedGoogle Scholar
  11. 11.
    Borgstahl GE, Parge HE, Hickey MJ, Beyer WF Jr, Hallewell RA, Tainer JA (1992) Cell 71:107–118CrossRefPubMedGoogle Scholar
  12. 12.
    Edwards RA, Whittaker MM, Whittaker JW, Jameson GB, Baker EN (1998) J Am Chem Soc 120:9684–9685CrossRefGoogle Scholar
  13. 13.
    Lah MS, Dixon MM, Pattridge KA, Stallings WC, Fee JA, Ludwig ML (1995) Biochemistry 34:1646–1660CrossRefPubMedGoogle Scholar
  14. 14.
    Guan Y, Hickey MJ, Borgstahl GEO, Hallewell RA, Lepock JR, O’Connor D, Hsieh Y, Nick HS, Silverman DN, Tainer JA (1998) Biochemistry 37:4722–4730CrossRefPubMedGoogle Scholar
  15. 15.
    Youn H-D, Kim E-J, Roe J-H, Hah YC, Kang S-O (1996) Biochem J 318:889–896PubMedGoogle Scholar
  16. 16.
    Youn H-D, Youn H, Lee J-W, Yim Y-I, Lee J-K, Hah YC, Kang S-O (1996) Arch Biochem Biophys 334:341–348CrossRefPubMedGoogle Scholar
  17. 17.
    Dupont CL, Neupane K, Shearer J, Palenik B (2008) Environ Microbiol 10:1831–1843CrossRefPubMedGoogle Scholar
  18. 18.
    Wuerges J, Lee J-W, Yim Y-I, Yim H-S, Kang S-O, Djinovic-Carugo K (2004) Proc Natl Acad Sci USA 101:8569–8574CrossRefPubMedGoogle Scholar
  19. 19.
    Barondeau DP, Kassmann CJ, Bruns CK, Tainer JA, Getzoff ED (2004) Biochemistry 43:8038–8047CrossRefPubMedGoogle Scholar
  20. 20.
    Choudhury SB, Lee J-W, Davidson G, Yim Y-I, Bose K, Sharma ML, Kang S-O, Cabelli DE, Maroney MJ (1999) Biochemistry 38:3744–3752CrossRefPubMedGoogle Scholar
  21. 21.
    Evans DJ (2005) Coord Chem Rev 249:1582–1595CrossRefGoogle Scholar
  22. 22.
    Kraulis PJ (1991) J Appl Crystallogr 24:946–950CrossRefGoogle Scholar
  23. 23.
    Merritt EA, Murphy MEP (1994) Acta Crystallogr D 50:869–873CrossRefPubMedGoogle Scholar
  24. 24.
    Fontecilla-Camps JC, Volbeda A, Cavazza C, Nicolet Y (2007) Chem Rev 107:4273–4303CrossRefPubMedGoogle Scholar
  25. 25.
    Werst MM, Davoust CE, Hoffman BM (1991) J Am Chem Soc 113:1533–1538CrossRefGoogle Scholar
  26. 26.
    Davoust CE, Doan PE, Hoffman BM (1996) J Magn Reson 119:38–44CrossRefGoogle Scholar
  27. 27.
    Schweiger A, Jeschke G (2001) Principles of pulse electron paramagnetic resonance. Oxford University Press, OxfordGoogle Scholar
  28. 28.
    Abragam A, Bleaney B (1986) Electron paramagnetic resonance of transition ions. Dover, New YorkGoogle Scholar
  29. 29.
    Hoffman BM, Martinsen J, Venters RA (1984) J Magn Reson 59:110–123Google Scholar
  30. 30.
    Hoffman BM, Venters RA, Martinsen J (1985) J Magn Reson 62:537–542Google Scholar
  31. 31.
    Hoffman BM, DeRose VJ, Doan PE, Gurbiel RJ, Houseman ALP, Telser J (1993) Biol Magn Reson 13:151–218Google Scholar
  32. 32.
    Doan PE (2003) In: Telser J (ed) Paramagnetic resonance of metallobiomolecules. American Chemical Society, Washington, pp 55–81CrossRefGoogle Scholar
  33. 33.
    Fan CL, Doan PE, Davoust CE, Hoffman BM (1992) J Magn Reson 98:62–72Google Scholar
  34. 34.
    Dikanov SA, Tsvetkov YD (1992) Electron spin echo envelope modulation (ESEEM) spectroscopy. CRC, Boca RatonGoogle Scholar
  35. 35.
    Mims WB (1984) J Magn Reson 59:291–306Google Scholar
  36. 36.
    Lee H-I, Doan PE, Hoffman BM (1999) J Magn Reson 140:91–107CrossRefPubMedGoogle Scholar
  37. 37.
    Lappin AG, Murray CK, Margerum DW (1978) Inorg Chem 17:1630–1634CrossRefGoogle Scholar
  38. 38.
    Lovecchio FV, Gore ES, Busch DH (1974) J Am Chem Soc 96:3109–3118CrossRefGoogle Scholar
  39. 39.
    Pinho D, Gomes P, Freire C, de Castro B (2001) Eur J Inorg Chem 2001(6):1483–1493CrossRefGoogle Scholar
  40. 40.
    Wang YL, Beach MW, Pappenhagen TL, Margerum DW (1988) Inorg Chem 27:4464–4472CrossRefGoogle Scholar
  41. 41.
    Seth J, Palaniappan V, Bocian DF (1995) Inorg Chem 34:2201–2206CrossRefGoogle Scholar
  42. 42.
    Brown TG, Hoffman BM (1980) Mol Phys 39:1073–1109CrossRefGoogle Scholar
  43. 43.
    Telser J, Fann Y-C, Renner MW, Fajer J, Wang S, Zhang H, Scott RA, Hoffman BM (1997) J Am Chem Soc 119:733–743CrossRefGoogle Scholar
  44. 44.
    Jiang F, McCracken J, Peisach J (1990) J Am Chem Soc 112:9035–9044CrossRefGoogle Scholar
  45. 45.
    Colaneri MJ, Peisach J (1992) J Am Chem Soc 114:5335–5341CrossRefGoogle Scholar
  46. 46.
    Mathews J, Walker RL (1965) Mathematical methods of physics. Benjamin, ElmsfordGoogle Scholar
  47. 47.
    True E, Nelson MJ, Venters RA, Orme-Johnson WH, Hoffman BM (1988) J Am Chem Soc 110:1935–1943CrossRefGoogle Scholar

Copyright information

© SBIC 2009

Authors and Affiliations

  1. 1.Department of ChemistryKyungpook National UniversityDaeguRepublic of Korea
  2. 2.Department of Life Science, Research Center for Natural SciencesHanyang UniversitySeoulRepublic of Korea
  3. 3.Laboratory of Biophysics, School of Biological Sciences, Institute of MicrobiologySeoul National UniversitySeoulRepublic of Korea
  4. 4.Department of ChemistryNorthwestern UniversityEvanstonUSA

Personalised recommendations