Metal-ion mutagenesis: Conversion of a purple acid phosphatase from sweet potato to a neutral phosphatase with the formation of an unprecedented catalytically competent MnIIMnII active site

Mitić, Nataša, Noble, Christopher J., Gahan, Lawrence R., Hanson, Graeme R. and Schenk, Gerhard (2009) Metal-ion mutagenesis: Conversion of a purple acid phosphatase from sweet potato to a neutral phosphatase with the formation of an unprecedented catalytically competent MnIIMnII active site. Journal of the American Chemical Society, 131 23: 8173-8179. doi:10.1021/ja900797u


Author Mitić, Nataša
Noble, Christopher J.
Gahan, Lawrence R.
Hanson, Graeme R.
Schenk, Gerhard
Title Metal-ion mutagenesis: Conversion of a purple acid phosphatase from sweet potato to a neutral phosphatase with the formation of an unprecedented catalytically competent MnIIMnII active site
Formatted title
Metal-ion mutagenesis: Conversion of a purple acid phosphatase from sweet potato to a neutral phosphatase with the formation of an unprecedented catalytically competent MnIIMnII active site
Journal name Journal of the American Chemical Society   Check publisher's open access policy
ISSN 0002-7863
1520-5126
1943-2984
Publication date 2009-06-17
Year available 2009
Sub-type Article (original research)
DOI 10.1021/ja900797u
Open Access Status
Volume 131
Issue 23
Start page 8173
End page 8179
Total pages 7
Editor Peter J. Stang
Place of publication Washington, DC, U.S.A.
Publisher American Chemical Society
Language eng
Subject C1
030201 Bioinorganic Chemistry
970103 Expanding Knowledge in the Chemical Sciences
Abstract The currently accepted paradigm is that the purple acid phosphatases (PAPs) require a heterovalent, dinuclear metal-ion center for catalysis. It is believed that this is an essential feature for these enzymes in order for them to operate under acidic conditions. A PAP from sweet potato is unusual in that it appears to have a specific requirement for manganese, forming a unique Fe-III-mu-(O)-Mn-II center under catalytically optimal conditions (Schenk et al. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 273). Herein, we demonstrate, with detailed electron paramagnetic resonance (EPR) spectroscopic and kinetic studies, that in this enzyme the chromophoric Fe-III can be replaced by Mn-II, forming a catalytically active, unprecedented antiferromagnetically coupled homodivalent Mn-II-mu-(H)OH-mu-carboxylato-Mn-II center in a PAP. However, although the enzyme is still active, it no longer functions as an acid phosphatase, having optimal activity at neutral pH. Thus, PAPs may have evolved from distantly related divalent dinuclear metallohydrolases that operate under pH neutral conditions by stabilization of a trivalent-divalent metal-ion core. The present Mn-II-Mn-II system models these distant relatives, and the results herein make a significant contribution to our understanding of the role of the chromophoric metal ion as an activator of the nucleophile. In addition, the detailed analysis of strain broadened EPR spectra from exchange-coupled dinuclear Mn-II-Mn-II centers described herein provides the basis for the full interpretation of the EPR spectra from other dinuclear Mn metalloenzymes.
Formatted abstract
The currently accepted paradigm is that the purple acid phosphatases (PAPs) require a heterovalent, dinuclear metal-ion center for catalysis. It is believed that this is an essential feature for these enzymes in order for them to operate under acidic conditions. A PAP from sweet potato is unusual in that it appears to have a specific requirement for manganese, forming a unique FeIII-μ-(O)-MnII center under catalytically optimal conditions (Schenk et al. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 273). Herein, we demonstrate, with detailed electron paramagnetic resonance (EPR) spectroscopic and kinetic studies, that in this enzyme the chromophoric FeIII can be replaced by MnII, forming a catalytically active, unprecedented antiferromagnetically coupled homodivalent MnII-μ-(H)OH-μ-carboxylato-MnII center in a PAP. However, although the enzyme is still active, it no longer functions as an acid phosphatase, having optimal activity at neutral pH. Thus, PAPs may have evolved from distantly related divalent dinuclear metallohydrolases that operate under pH neutral conditions by stabilization of a trivalent−divalent metal-ion core. The present MnII−MnII system models these distant relatives, and the results herein make a significant contribution to our understanding of the role of the chromophoric metal ion as an activator of the nucleophile. In addition, the detailed analysis of strain broadened EPR spectra from exchange-coupled dinuclear MnII−MnII centers described herein provides the basis for the full interpretation of the EPR spectra from other dinuclear Mn metalloenzymes.
Keyword Pig allantoic fluid
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

 
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Created: Thu, 03 Sep 2009, 17:52:59 EST by Mr Andrew Martlew on behalf of School of Chemistry & Molecular Biosciences