Abstract
A protein histidine phosphatase (PHP) from vertebrates was first identified in 2002. Here we describe the expression of that PHP in Escherichia coli and purification of the recombinant protein. In addition, a detailed protocol is provided describing determination of PHP activity in vitro. Proteins phosphorylated on histidine residues in general cannot be easily obtained. This also applies to the substrates of PHP. To circumvent that obstacle, assay conditions are introduced enabling scientists to study PHP activity using a substrate within crude homogenates of cells and tissues.
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Boyer, P. D., DeLuca, M., Ebner, K. E., Hultquist, D. E., and Peter, J. B. (1962) Identification of phospohistidine in digests from a probable intermediate of oxidative phosphorylation. J. Biol. Chem. 237, PC3306–PC3308.
Mitchell, R. A., Butler, L. G., and Boyer, P.D. (1964) The association of readily-soluble bound phosphohistidine from mitochondria with succinate thiokinase. Biochem. Biophys. Res. Commun. 16, 545–550.
Walinder, O. (1968) Identification of a phosphate-incorporating protein from bovine liver as nucleoside diphosphate kinase and isolation of 1-32P-phosphohistidine, 3-32P-phosphohistidine, and N-ε-32P-phospholysine from erythrocytic nucleoside diphosphate kinase, incubated with adenosine triphosphate-32P. J. Biol. Chem. 243, 3947–3952.
Feldman, F. and Butler, L. G. (1969) Detection and characterization of the phosphorylated form of microsomal glucose-6-phosphatase. Biochem. Biophys. Res. Commun. 36, 119–125.
Cottam, G. L., Srere, P. A. (1969) Nature of the phosphorylated residue in citrate clevage enzyme. Biochem. Biophys. Res. Commun. 35, 895–900.
Wieland, T., Nürnberg, B., Ulibarri, I., Kaldenberg-Stasch, S., Schultz, G., and Jakobs, K. H. (1993) Guanine nucleotide-specific phosphate transfer by guanine nucleotide-binding regulatory protein beta-subunits. Characterization of the phosphorylated amino acid. J. Biol. Chem. 268, 18,111–18,118.
Crovello, C. S., Furie, B. C., and Furie, B. (1995) Histidine phosphorylation of P-selectin upon stimulation of human platelets: a novel pathway for activation-dependent signal transduction. Cell 82, 279–286.
Muimo, R., Hornickova, Z., Riemen, C. E., Gerke, V., Matthews, H., and Mehta, A. (2000) Histidine phosphorylation of annexin I in airway epithelia. J. Biol. Chem. 275, 36,632–36,636.
Alex, L. A. and Simon, M. I. (1994) Protein histidine kinases and signal transduction in prokaryotes and eukaryotes. Trends Genet. 10, 133–138.
Klumpp, S., Hermesmeier, J., Selke, D., et al. (2002) Protein histidine phosphatase: a novel enzyme with potency for neuronal signaling. J. Cereb. Blood. Flow Metab. 22, 1420–1424.
Ek, P., Pettersson, G., Ek, B., Gong, F., Li, J-P., and Zetterquist, Ö. (2002) Identification and characterization of mammalian 14-kDa phosphohistidine phosphatase. Eur. J. Biochem. 269, 5016–5023.
Klumpp, S., Bechmann, G., Mäurer, A., Selke, D., and Krieglstein, J. (2003) ATP-citrate lyase as a substrate of protein histidine phosphatase in vertebrates. Biochem. Biophys. Res. Commun. 306, 110–115.
Mäurer, A., Wieland, T., Meissl, F., et al. (2005) The β-subunit of G proteins is a substrate of protein histidine phosphatase. Biochem. Biophys. Res. Commun. 334, 1115–1120.
Motojima, K. and Goto, S. (1993) A protein histidine kinase induced in rat liver by peroxisome proliferators. In vitro activation by Ras protein and guanine nucleotides. FEBS Lett. 319, 75–79.
Besant, P. G., Tan, E., and Attwood, P. V. (2003) Mammalian protein histidine kinases. Int. J. Biochem. Cell Biol. 35, 297–309.
Noiman, S. and Shaul, Y. (1995) Detection of histidine-phospho-proteins in animal tissues. FEBS Lett. 364, 63–66.
Baneyx, F. and Mujacic, M. (2004) Recombinant protein folding and misfolding in Escherichia coli. Nature Biotechnol. 22, 1399–1408.
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© 2007 Humana Press Inc.
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Bäumer, N., Mäurer, A., Krieglstein, J., Klumpp, S. (2007). Expression of Protein Histidine Phosphatase in Escherichia coli, Purification, and Determination of Enzyme Activity. In: Moorhead, G. (eds) Protein Phosphatase Protocols. Methods in Molecular Biology, vol 365. Springer, Totowa, NJ. https://doi.org/10.1385/1-59745-267-X:247
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DOI: https://doi.org/10.1385/1-59745-267-X:247
Publisher Name: Springer, Totowa, NJ
Print ISBN: 978-1-58829-711-2
Online ISBN: 978-1-59745-267-0
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