Chapter 14 Protein Histidine Phosphorylation

This chapter discusses protein histidine phosphorylation, protein histidine kinases (HKs), and protein histidine phosphatases (PHPs) and describes their roles in bacteria, fungi, plants, and mammalian cells. Current methods of detection of phosphohistidine in proteins are also described, including HK assays, phosphoamino acid analysis, and approaches involving mass spectrometric (MS) methods. The HKs in bacteria, fungi, and plants are two-component protein systems composed of two major functional parts: the HK and the response regulator protein. The receptor or sensor protein that has the HK activity exists in the cell membrane as a preformed dimer or in some cases, may dimerize in response to the extracellular signal. One of the simplest ways to confirm that the site of phosphorylation in a phosphoprotein is a particular amino acid is to perform phosphoamino acid analysis. In this process, the protein substrate is phosphorylated using a nucleotide in which the γ-phosphate is radiolabelled, commonly with 32P, resulting in the formation of a [32P] phosphoprotein product. Phosphohistidine in proteins is directly detected using 31P and 1H nuclear magnetic resonance (NMR).