The structure and function of HPr.

Histidine-containing phosphocarrier protein, HPr, was one of the early protein tertiary structures determined by two-dimensional 1H-NMR. Tertiary structures for HPrs from Escherichia coli, Bacillus subtilis, and Staphylococcus aureus have been obtained by 1H NMR and the overall folding pattern of HPr is highly conserved, a betaalpha betabeta alphabeta alpha arrangement of three alpha-helices overlaying a four-stranded beta-sheet. High-resolution structures for HPrs from E. coli and B. subtilis have been obtained using 15N- and 13C-labeled proteins. The first application of NMR to the understanding of the structure and function of HPr was to describe the phosphohistidine isomer, Ndelta1-P-histidine in S. aureus phospho-HPr, and the unusual pKas of the His-15 side chain. The pKa values for the His-15 imidazole from more recent studies are 5.4 for HPr and 7.8 for phospho-HPr from E. coli, for example. A consensus description of the active site is proposed for HPr and phospho-HPr. In HPr, His-15 has a defined conformation and N-caps helix A, and is thus affected by the helix dipole. His-15 undergoes a small conformational change upon phosphorylation, a movement to allow the phosphoryl group to be positioned such that it forms hydrogen bonds with the main chain amide nitrogens of residue 16 (not conserved) and Arg-17. Interactions between residue 12 side chain (not conserved: asparagine, serine, and threonine) and His-15, and between the Arg-17 guanidinium group and the phosphoryl group, are either weak or transitory.