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Highly efficient precipitation of phosphoproteins using trivalent europium, terbium, and erbium ions

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Abstract

This study describes a highly efficient method for the selective precipitation of phosphoproteins by trivalent europium, terbium, and erbium metal ions. These metal cations belong to the group of lanthanides and are known to be hard acceptors with an overwhelming preference for oxygen-containing anions such as phosphates to which they form very tight ionic bonds. The method could be successfully applied to specifically precipitate phosphoproteins from complex samples including milk and egg white by forming solid metal–protein complexes. Owing to the low solubility product of the investigated lanthanide salts, the produced metal–protein complexes showed high stability. The protein pellets were extensively washed to remove nonphosphorylated proteins and contaminants. For the analysis of proteins the pellets were first dissolved in 30 % formic acid and subjected to matrix-assisted laser desorption/ionization–time of flight (MALDI-TOF) MS. For peptide mass-fingerprint analysis the precipitated phosphoproteins were enzymatically digested using microwave-assisted digestion. The method was found to be highly specific for the isolation and purification of phosphoproteins. Protein quantification was performed by colorimetric detection of total precipitated phosphoproteins and revealed more than 95 % protein recovery for each lanthanide salt.

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Acknowledgements

This work was supported by the Austrian Science Foundation (FWF), SFB-Project 021 (Vienna, Austria).

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Authors and Affiliations

  1. Institute of Analytical Chemistry and Radiochemistry, Leopold-Franzens University, 6020, Innsbruck, Austria

    Yüksel Güzel, Matthias Rainer, Munazza Raza Mirza & Günther K. Bonn

  2. Institute of Analytical Chemistry and Radiochemistry, Leopold-Franzens University, Innsbruck, Innrain 52a, 6020, Innsbruck, Austria

    Matthias Rainer

Authors
  1. Yüksel Güzel
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  2. Matthias Rainer
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  3. Munazza Raza Mirza
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  4. Günther K. Bonn
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Correspondence to Matthias Rainer.

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Güzel, Y., Rainer, M., Mirza, M.R. et al. Highly efficient precipitation of phosphoproteins using trivalent europium, terbium, and erbium ions. Anal Bioanal Chem 403, 1323–1331 (2012). https://doi.org/10.1007/s00216-012-5917-7

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  • DOI: https://doi.org/10.1007/s00216-012-5917-7

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