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    Probing the role of the divalent metal ion in uteroferrin using metal ion replacement and a comparison to isostructural biomimetics

    Schenk, Gerhard and Peralta, Rosely A. and Batista, Suzana Cimara and Bortoluzzi, Adailton J. and Szpoganicz, Bruno and Dick, Andrew J. and Herrald, Paul and Hanson, Graeme R. and Szilagyi, Robert K. and Riley, Mark J. and Gahan, Lawrence R. and Neves, Ademir (2008) Probing the role of the divalent metal ion in uteroferrin using metal ion replacement and a comparison to isostructural biomimetics. Journal of Biological Inorganic Chemistry, 13 (1). pp. 139-155. ISSN 0949-8257

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    Purple acid phosphatases (PAPs) are a group of heterovalent binuclear metalloenzymes that catalyze the hydrolysis of phosphomonoesters at acidic to neutral pH. While the metal ions are essential for catalysis, their precise roles are not fully understood. Here, the Fe(III)Ni(II) derivative of pig PAP (uteroferrin) was generated and its properties were compared with those of the native Fe(III)Fe(II) enzyme. The kcat of the Fe(III)Ni(II) derivative (approximately 60 s–1) is approximately 20% of that of native uteroferrin, and the Ni(II) uptake is considerably faster than the reconstitution of full enzymatic activity, suggesting a slow conformational change is required to attain optimal reactivity. An analysis of the pH dependence of the catalytic properties of Fe(III)Ni(II) uteroferrin indicates that the l-hydroxide is the likely nucleophile. Thus, the Ni(II) derivative employs a mechanism similar to that proposed for the Ga(III)Zn(II) derivative of uteroferrin, but different from that of the native enzyme, which uses a terminal Fe(III)-bound nucleophile to initiate catalysis. Binuclear Fe(III)Ni(II) biomimetics with coordination environments similar to the coordination environment of uteroferrin were generated to provide both experimental benchmarks (structural and spectroscopic) and further insight into the catalytic mechanism of hydrolysis. The data are consistent with a reaction mechanism employing an Fe(III)-bound terminal hydroxide as a nucleophile, similar to that proposed for native uteroferrin and various related isostructural biomimetics. Thus, only in the uteroferrin- catalyzed reaction are the precise details of the catalytic mechanism sensitive to the metal ion composition, illustrating the significance of the dynamic ligand environment in the protein active site for the optimization of the catalytic efficiency.

    Item Type: Article
    Keywords: Binuclear metallohydrolases; Purple acid phosphatases;  Uteroferrin;  Catalysis; Metal ion replacement;
    Academic Unit: Faculty of Science and Engineering > Chemistry
    Item ID: 3693
    Depositing User: Gary Schenk
    Date Deposited: 29 May 2012 09:02
    Journal or Publication Title: Journal of Biological Inorganic Chemistry
    Publisher: Springer Verlag
    Refereed: Yes
    Use Licence: This item is available under a Creative Commons Attribution Non Commercial Share Alike Licence (CC BY-NC-SA). Details of this licence are available here

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