Phosphate ester cleavage promoted by a tetrameric iron(III) complex
- 198 Downloads
The purple acid phosphatases (PAPs) are the only binuclear metallohydrolases where the necessity for a heterovalent active site [Fe(III)–M(II) (M is Fe, Zn or Mn)] for catalysis has been established. The paradigm for the construction of PAP biomimetics, both structural and functional, is that the ligands possess characteristics which mimic those of the donor sites of the metalloenzyme and permit discrimination between trivalent and divalent metal ions. The donor atom set of the ligand 2-((2-hydroxy-5-methyl-3-((pyridin-2-ylmethylamino)methyl)benzyl)(2-hydroxybenzyl)amino)acetic acid (H3HPBA) mimics that of the active site of PAP although the iron(III) complex of this ligand has been characterized as the tetramer [Fe4(HPBA)2(μ-CH3COO)2(μ-O)(μ-OH)(OH2)2]ClO4·5H2O. The phosphoesterase-like activity of the complex in 1:1 acetonitrile/water has now been investigated using the substrate 2,4-bis(dinitrophenyl)phosphate. The pH dependence of the catalytic rate revealed a non-symmetric bell-shaped profile, with a finite but non-zero rate at high pH. Unlike the traditional approach usually employed to analyse these bell-shaped profiles, the approach used here involved incorporating additional species which contribute to the overall activity. Employing this approach, we show that the complex has a k cat of 1.6 (±0.2) × 10−3 s−1, three kinetically relevant pK a values of 5.3, 6.2 and 8.4, with K M of 7.4 ± 0.6 mM. The kinetic parameters are similar to those reported for heterovalent PAP biomimetics. Additionally, it is observed that, unlike the enzyme, the oxidation state is not the determining factor for catalytic activity.
KeywordsPurple acid phosphatase Phosphodiester-degrading enzyme Binuclear metallohydrolases Iron complexes Biomimetics
This work was funded by a grant from the Australian Research Council (DP0986292).
- 2.Klabunde T, Krebs B (1997) Struct Bonding (Berlin) 89:177–198Google Scholar
- 15.Boudalis AK, Aston RE, Smith SJ, Mirams RE, Riley MJ, Schenk G, Blackman AG, Hanton LR, Gahan LR (2007) Dalton Trans 5132–5139Google Scholar
- 27.Systat Software (2006) SigmaPlot for Windows, version 10. Systat Software, San JoseGoogle Scholar
- 28.Binstead RA, Zuberbuhler AD (1993–1995) SPECFIT: a program for global least squares fitting of equilibria and kinetics systems using factor analysis and Marquardt minimization, version 2.09. Spectrum Software Associates, Chapel HillGoogle Scholar
- 32.Purich DL, Allison RD (2000) Handbook of biochemical kinetics. Academic Press, San Diego, p 544Google Scholar
- 33.von Euler H, Josephson K, Myrback K (1924) Z Physiol Chem 134:39–49Google Scholar
- 34.Segel IH (1975) Enzyme kinetics: behavior and analysis of rapid equilibrium and steady-state enzyme systems. Wiley-Interscience, New YorkGoogle Scholar
- 38.Li D-F, Liao Z-R, Wei Y-G, Wang M, Chen W-X, Li W-K, Mao X-A (2003) Dalton Trans 2164–2169Google Scholar
- 39.Sung N, Kim T-Y (2006) Agric Chem Biotechnol 49:86–89Google Scholar
- 42.Tanaka L, Yamada S (1976) J Chem Soc Chem Commun 178–179Google Scholar