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The reaction mechanism of the Ga(III)Zn(II) derivative of uteroferrin and corresponding biomimetics

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Abstract

Purple acid phosphatase from pig uterine fluid (uteroferrin), a representative of the diverse family of binuclear metallohydrolases, requires a heterovalent Fe(III)Fe(II) center for catalytic activity. The active-site structure and reaction mechanism of this enzyme were probed with a combination of methods including metal ion replacement and biomimetic studies. Specifically, the asymmetric ligand 2-bis{[(2-pyridylmethyl)-aminomethyl]-6-[(2-hydroxybenzyl)(2-pyridylmethyl)]aminomethyl}-4-methylphenol and two symmetric analogues that contain the softer and harder sites of the asymmetric unit were employed to assess the site selectivity of the trivalent and divalent metal ions using 71Ga NMR, mass spectrometry and X-ray crystallography. An exclusive preference of the harder site of the asymmetric ligand for the trivalent metal ion was observed. Comparison of the reactivities of the biomimetics with Ga(III)Zn(II) and Fe(III)Zn(II) centers indicates a higher turnover for the former, suggesting that the M(III)-bound hydroxide acts as the reaction-initiating nucleophile. Catalytically active Ga(III)Zn(II) and Fe(III)Zn(II) derivatives were also generated in the active site of uteroferrin. As in the case of the biomimetics, the Ga(III) derivative has increased reactivity, and a comparison of the pH dependence of the catalytic parameters of native uteroferrin and its metal ion derivatives supports a flexible mechanistic strategy whereby both the μ-(hydr)oxide and the terminal M(III)-bound hydroxide can act as nucleophiles, depending on the metal ion composition, the geometry of the second coordination sphere and the substrate.

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Notes

  1. The more alkaline pH optimum for bovine PAP may be consistent with a number of other catalytic variations (i.e., k cat, substrate and fluoride inhibition) observed between these otherwise homologous proteins. The origin of these differences is not yet fully understood but may be associated with a mobile loop in the vicinity of the binuclear site [1, 22–28].

  2. The obvious caveat of this assignment is that the previously mentioned ENDOR study [21] demonstrated that no terminal water ligand may be present in resting Uf. These conflicting data may be reconciled if the addition of substrate to Uf leads to the coordination of an Fe(III)-bound water molecule (note that in the structure of PAP from rat, electron density ascribed to a terminal Fe(III)-bound water molecule is observed in the presence of the bound substrate mimic sulfate [18]). Alternatively, pK es2 may describe the deprotonation of the μ-hydroxide. However, in this case the substitution of Fe(II) by Zn(II) would be expected to have a greater effect on pK es2. Furthermore, the exchange coupling determined from magnetic susceptibility measurements [66] indicates that at pH 4.90 a μ-hydroxide is present in native Fe(III)Fe(II) Uf.

  3. Note, however, that at least in bovine PAP the latter is not corroborated by the apparent lack of burst kinetics [24] (see also footnote 1).

References

  1. Mitić N, Smith SJ, Neves A, Guddat LW, Gahan LR, Schenk G (2006) Chem Rev 106:3338–3363

    Article  PubMed  Google Scholar 

  2. Mitić N, Schenk G, Hanson GR (2007) In: Hanson GR, Berliner LJ (eds) Biological magnetic resonance: high resolution EPR: applications to metalloenzymes and metals in medicine, vol 28. Springer, New York (in press)

  3. Klabunde T, Krebs B (1997) Struct Bonding 89:177–198

    CAS  Google Scholar 

  4. Oddie GW, Schenk G, Angel NZ, Walsh N, Guddat LW, de Jersey J, Cassady AI, Hamilton SE, Hume DA (2000) Bone 27:575–584

    Article  PubMed  CAS  Google Scholar 

  5. Schenk G, Guddat LW, Ge Y, Carrington LE, Hume DA, Hamilton S, de Jersey J (2000) Gene 250:117–125

    Article  PubMed  CAS  Google Scholar 

  6. Wang DL, Holz RC, David SS, Que L, Stankovich MT (1991) Biochemistry 30:8187–8194

    Article  PubMed  CAS  Google Scholar 

  7. Bernhardt PV, Schenk G, Wilson GJ (2004) Biochemistry 43:10387–10392

    Article  PubMed  CAS  Google Scholar 

  8. Beck JL, McConachie LA, Summors AC, Arnold WN, de Jersey J, Zerner B (1986) Biochim Biophys Acta 869:61–68

    CAS  Google Scholar 

  9. Durmus A, Eicken C, Sift BH, Kratel A, Kappi R, Hütterman J, Krebs B (1999) Eur J Biochem 260:709–716

    Article  PubMed  CAS  Google Scholar 

  10. Schenk G, Ge Y, Carrington LE, Wynne CJ, Searle IR, Carroll BJ, Hamilton S, de Jersey J (1999) Arch Biochem Biophys 370:183–189

    Article  PubMed  CAS  Google Scholar 

  11. Schenk G, Boutchard CL, Carrington LE, Noble CJ, Moubaraki B, Murray KS, de Jersey J, Hanson GR, Hamilton S (2001) J Biol Chem 276:19084–19088

    Article  PubMed  CAS  Google Scholar 

  12. Antanaitis BC, Aisen P, Lilienthal HR (1983) J Biol Chem 258:3166–3172

    PubMed  CAS  Google Scholar 

  13. Averill BA, Davis JC, Burman S, Zirino T, Sanders-Loehr J, Loehr TM, Sage JT, Debrunner PG (1987) J Am Chem Soc 109:3760–3767

    Article  CAS  Google Scholar 

  14. Yang Y-S, McCormick JM, Solomon EI (1997) J Am Chem Soc 119:11832–11842

    Article  CAS  Google Scholar 

  15. Klabunde T, Sträter N, Fröhlich R, Witzel H, Krebs B (1996) J Mol Biol 259:737–748

    Article  PubMed  CAS  Google Scholar 

  16. Guddat LW, McAlpine AS, Hume D, Hamilton S, de Jersey J, Martin JL (1999) Structure 7:757–767

    Article  PubMed  CAS  Google Scholar 

  17. Schenk G, Gahan LR, Carrington LE, Mitić N, Valizadeh M, Hamilton SE, de Jersey J, Guddat LW (2005) Proc Natl Acad Sci USA 102:273–278

    Article  PubMed  CAS  Google Scholar 

  18. Lindqvist Y, Johansson E, Kaija H, Vihko P, Schneider G (1999) J Mol Biol 291:135–147

    Article  PubMed  CAS  Google Scholar 

  19. Sträter N, Jasper B, Scholte M, Krebs B, Duff AP, Langley DB, Han R, Averill BA, Freeman HC, Guss JM (2005) J Mol Biol 351:233–246

    Article  PubMed  Google Scholar 

  20. Sträter N, Klabunde T, Tucker P, Witzel H, Krebs B (1995) Science 268:1489–1492

    Article  PubMed  Google Scholar 

  21. Smoukov SK, Quaroni L, Wang X, Doan PE, Hoffman BM, Que L Jr (2002) J Am Chem Soc 124:2595–2603

    Article  PubMed  CAS  Google Scholar 

  22. Valizadeh M, Schenk G, Nash K, Oddie GW, Guddat LW, Hume DA, de Jersey J, Burke J, Terrence R., Hamilton S (2004) Arch Biochem Biophys 424:154–162

    Article  PubMed  CAS  Google Scholar 

  23. Twitchett MB, Schenk G, Aquino MAS, Yiu DTY, Lau T-C, Sykes AG (2002) Inorg Chem 41:5787–5794

    Article  PubMed  CAS  Google Scholar 

  24. Merkx M, Pinkse MWH, Averill BA (1999) Biochemistry 38:9914–9925

    Article  PubMed  CAS  Google Scholar 

  25. Aquino MAS, Lim J-S, Sykes AG (1994) J Chem Soc Dalton Trans 429–436

  26. Merkx M, Averill BA (1998) Biochemistry 37:8490–8497

    Article  PubMed  CAS  Google Scholar 

  27. Merkx M, Averill BA (1999) J Am Chem Soc 121:6683–6689

    Article  CAS  Google Scholar 

  28. Mitić N, Valizadeh M, Leung EWW, de Jersey J, Hamilton S, Hume DA, Cassady AI, Schenk G (2005) Arch Biochem Biophys 439:154–464

    Article  PubMed  Google Scholar 

  29. Wang X, Ho RYN, Whiting AK, Que L Jr (1999) J Am Chem Soc 121:9235–9236

    Article  CAS  Google Scholar 

  30. Aramini JM, McIntyre DD, Vogel HJ (1994) J Am Chem Soc 116:11506–11511

    Article  CAS  Google Scholar 

  31. Vo E, Wang HC, Germanas JP (1997) J Am Chem Soc 119:1934–1940

    Article  CAS  Google Scholar 

  32. Johnson KA, Brereton PS, Verhagen MFJM, Calzolai L, La Mar GN, Adams MWW, Amster IJ (2001) J Am Chem Soc 123:7935–7936

    Article  PubMed  CAS  Google Scholar 

  33. Maher M, Cross M, Wilce MCJ, Guss JM, Wedd AG (2004) Acta Crystallogr Sect D 60:298–303

    Article  Google Scholar 

  34. Woodworth RC, Butcher ND, Brown SA, Brown-Mason A (1987) Biochemistry 26:3115–3120

    Article  PubMed  CAS  Google Scholar 

  35. Ueno T, Kousumi Y, Yoshizawa-Kumagaye K, Nakajima K, Ueyama N, Okamura T, Nakamura A (1998) J Am Chem Soc 120:12264–12273

    Article  CAS  Google Scholar 

  36. Borovik AS, Papaefthymiou V, Taylor LF, Anderson OP, Que L (1989) J Am Chem Soc 111:6183–6195

    Article  CAS  Google Scholar 

  37. Neves A, de Brito MA, Drago V, Griesar K, Haase W (1995) Inorg Chim Acta 237:131–135

    Article  CAS  Google Scholar 

  38. Lambert E, Chabut B, Chardon-Noblat S, Deronzier A, Chottard G, Bousseksou A, Tuchagues J-P, Laugier J, Bardet M, Latour J-M (1997) J Am Chem Soc 119:9424–9437

    Article  CAS  Google Scholar 

  39. Batista SC, Neves A, Bortoluzzi AJ, Vencato I, Peralta RA, Szpoganicz B, Aires VVE, Terenzi H, Severino PC (2003) Inorg Chem Commun 6:1161–1165

    Article  CAS  Google Scholar 

  40. Karsten P, Neves A, Bortoluzzi AJ, Strahle J, Maichle-Mossmer C (2002) Inorg Chem Commun 5:434–438

    Article  CAS  Google Scholar 

  41. Karsten P, Neves A, Bortoluzzi AJ, Lanznaster M, Drago V (2002) Inorg Chem 41:4624–4626

    Article  PubMed  CAS  Google Scholar 

  42. Lanznaster M, Neves A, Bortoluzzi AJ, Szpoganicz B, Schwingel E (2002) Inorg Chem 41:5641–5643

    Article  PubMed  CAS  Google Scholar 

  43. Lanznaster M, Neves A, Bortoluzzi AJ, Aires VVE, Szpoganicz B, Terenzi H, Severino PC, Fuller JM, Drew SC, Gahan LR, Hanson GR, Riley MJ, Schenk G (2005) J Biol Inorg Chem 10:319–332

    Article  PubMed  CAS  Google Scholar 

  44. Neves A, Lanznaster M, Bortoluzzi AJ, Peralta RA, Casellato A, Castellano EE, Herrald P, Riley MJ, Schenk G (2007) J Am Chem Soc 129:7486–7487

    Article  PubMed  CAS  Google Scholar 

  45. Torelli S, Belle C, Gautier-Luneau I, Pierre J-L (2000) Inorg Chem 39:3526–3536

    Article  PubMed  CAS  Google Scholar 

  46. Krebs B, Schepers K, Bremer B, Henkel G, Althaus E, Muller-Warmuth W, Griesar K, Haase W (1994) Inorg Chem 33:1907–1914

    Article  CAS  Google Scholar 

  47. Martell AE, Montekaitis RJ (1992) Determination and use of stability constants, 2nd edn. VCD, New York

    Google Scholar 

  48. Spek AL (1997) PLATON: molecular geometry and plotting program. University of Utrecht, The Netherlands

    Google Scholar 

  49. North AC, Phillips DC, Matthews FS (1968) Acta Crystallogr Sect A 24:351–359

    Article  Google Scholar 

  50. Sheldrick GM (1990) SHELXS97: program for the solution of crystal structures. University of Göttingen, Germany

    Google Scholar 

  51. Sheldrick GM (1997) SHELXL97: program for the refinement of crystal structures. University of Göttingen, Germany

    Google Scholar 

  52. Campbell HD, Dionysius DA, Keough DT, Wilson BE, de Jersey J, Zerner B (1978) Biochem Biophys Res Commun 82:615–620

    Article  PubMed  CAS  Google Scholar 

  53. Ravel B, Newville M (2005) J Synchrotron Radiat 12:537–541

    Article  PubMed  CAS  Google Scholar 

  54. Delpuech JJ (1983) In: Laszlo P (ed) Aluminium-27. Academic, New York, pp 153–195

  55. Andre JP, Macke HR (2003) J Inorg Biochem 97:315–323

    Article  PubMed  CAS  Google Scholar 

  56. Caravan P, Orvig C (1997) Inorg Chem 36:236–248

    Article  CAS  Google Scholar 

  57. Holman TR, Juarez-Garcia C, Hendrich MP, Que L Jr, Munck E (1990) J Am Chem Soc 112:7611–7618

    Article  CAS  Google Scholar 

  58. Torelli S, Belle C, Gautier-Luneau I, Hammam S, Pierre J-L (2002) Inorg Chim Acta 144–147

  59. Segel IH (1975) Enzyme kinetics: behavior and analysis of rapid equilibrium and steady-state enzyme systems. Wiley, New York

    Google Scholar 

  60. Twitchett MB, Sykes AG (1999) Eur J Inorg Chem 2105–2115

  61. Beck JL, Keough DT, de Jersey J, Zerner B (1984) Biochim Biophys Acta 791:357–363

    PubMed  CAS  Google Scholar 

  62. Wang X, Que L (1998) Biochemistry 37:7813–7821

    Article  PubMed  CAS  Google Scholar 

  63. Wang X, Randall CR, True AE, Que L Jr (1996) Biochemistry 35:13946–13954

    Article  PubMed  CAS  Google Scholar 

  64. Schenk G, Carrington LE, Hamilton SE, de Jersey J, Guddat LW (1999) Acta Crystallogr Sect D 55:2051–2052

    Article  CAS  Google Scholar 

  65. Funhoff EG, Wang Y, Andersson G, Averill BA (2005) FEBS J 272:2968–2977

    Article  PubMed  CAS  Google Scholar 

  66. Day E, David S, Peterson J, Dunham W, Bonvoisin J, Sands R, Que L Jr (1988) J Biol Chem 263:15561–15567

    PubMed  CAS  Google Scholar 

  67. Pinkse MWH, Merkx M, Averill BA (1999) Biochemistry 38:9926–9936

    Article  PubMed  CAS  Google Scholar 

  68. Dikiy A, Funhoff EG, Averill BA, Ciuril S (2002) J Am Chem Soc 124:13974–13975

    Article  PubMed  CAS  Google Scholar 

  69. Funhoff EG, de Jongh TE, Averill BA (2005) J Biol Inorg Chem 10:550–563

    Article  PubMed  CAS  Google Scholar 

  70. Elliott TW, Mitić N, Gahan LR, Guddat LW, Schenk G (2006) J Braz Chem Soc 17:1558–1565

    Article  CAS  Google Scholar 

  71. Cox RS, Schenk G, Mitić N, Gahan LR, Hengge AC (2007) J Am Chem Soc 129:9550–9551

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

This work was funded by a grant from the Australian Research Council (DP0558652), and CNPq and FAPESC from Brazil. X-ray absorption spectroscopy data collection was performed at the Australian National Beamline Facility (ANBF), Tsukuba, Japan, with support from the Australian Synchrotron Research Program, funded by the Commonwealth of Australia under the Major National Research Facilities Program. We also thank G. Foran for help in data collection.

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Authors and Affiliations

  1. School of Molecular and Microbial Sciences, The University of Queensland, St Lucia, QLD, 4072, Australia

    Sarah J. Smith, Annelise Casellato, Kieran S. Hadler, Nataša Mitić, Mark J. Riley, Gerhard Schenk & Lawrence R. Gahan

  2. Departamento de Química, Universidade Federal de Santa Catarina, Florianópolis, SC, 88040-900, Brazil

    Annelise Casellato, Adailton J. Bortoluzzi, Bruno Szpoganicz & Ademir Neves

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  1. Sarah J. Smith
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Correspondence to Gerhard Schenk or Ademir Neves.

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Smith, S.J., Casellato, A., Hadler, K.S. et al. The reaction mechanism of the Ga(III)Zn(II) derivative of uteroferrin and corresponding biomimetics. J Biol Inorg Chem 12, 1207–1220 (2007). https://doi.org/10.1007/s00775-007-0286-y

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