Abstract

The crystal structure of the histidine-containing phosphocarrier protein (HPr) of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) from Bacillus subtilis has been determined at 2.0-A resolution and refined to a crystallographic residual error R-factor of 0.150. The secondary-structure folding topology of the molecule is that of an open-face beta-sandwich formed by four antiparallel beta-strands packed against three alpha-helices. The active-site histidine, His-15, caps the N terminus of the first helix, suggesting that the helix dipole plays a role in stabilizing the phosphorylated state of the histidine. A sulfate anion located between His-15 and the neighboring Arg-17 has been identified in the electron-density map. Association of this negatively charged species with the two key catalytic residues implies that the crystal structure resembles the phosphorylated state of the protein. A model of the phosphorylated form of the molecule is proposed, in which the negatively charged phosphoryl group interacts with two main-chain nitrogen atoms of the following helix and with the guanidinium group of Arg-17. It is also proposed that the phosphoryl transfer from HPr to the IIA domain of the glucose permease involves Arg-17 switching between two salt bridges: one with the phosphorylated histidyl of HPr and the other with two aspartyl residues associated with the active site of the IIA domain of glucose permease, which are accessible upon complex formation.