Studies on Phosphohistidine Phosphatase 1 : What? Where? Why?

Abstract: Phosphohistidine phosphatase 1 (PHPT1) is a small protein, consisting of 125 amino acids, that catalyzes the dephosphorylation of histidine but does not have any activity towards other phosphorylated amino acids. PHPT1 was identified in 2002, and is so far the only mammalian histidine phosphatase known, but still little is known about its physiological role. No mammalian histidine kinases have hitherto been identified.Phosphorylation is one of the most important ways in which the structure and activity of a protein may be changed after translation. Proteins are phosphorylated on the side chain of amino acid residues. When a hydroxyl is phosphorylated the result is a phosphoester and when a nitrogen is phosphorylated the result is a phosphoamidate. Histidine may be phosphorylated on either of the two nitrogens of the imidazole ring of the side chain. The resulting phosphoamidate bond is labile and rich in energy, which makes histidine phosphorylation highly reversible and flexible. However, histidine phosphorylation is less studied than that of the phosphoesters due to the acid lability of the phosphoamidate bond.The work described in this thesis was focused on further elucidating the physiological role of PHPT1. Amino acid residues of importance for the activity of PHPT1 were identified, and mutants with decreased phosphatase activity were produced. These mutants have been used in studies on the function of PHPT1. By using immunohistochemical methodology the localization of PHPT1 in both mouse and human tissues was determined, with mainly similar results. A general finding was that expression of PHPT1 was high in epithelial cells with short turnover time, indicating that PHPT1 may have an important role in proliferating cells. We have also developed a comparatively fast and simple screening method for determination of PHPT1 activity. Since research in this field has been hampered by the lack of efficient and practical methodology, hopefully this new method will be an asset in search of inhibitors for PHPT1, which in turn may be used for detection of the elusive mammalian histidine kinases, the finding of which may give major breakthroughs in the field.