STRUCTURE OF THE IIA-DOMAIN OF THE GLUCOSE PERMEASE OF BACILLUS-SUBTILIS AT 2.2-A RESOLUTION

The crystal structure of the IIA domain of the glucose permease of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) from Bacillus subtilis has been determined at 2.2-angstrom resolution. Refinement of the structure is in progress, and the current R-factor is 0.201 (R = SIGMA-h parallel-to F(o)\ - \F(c) parallel-to/SIGMA-h\F(o)\, where \F(o)\ and \F(c)\ are the observed and calculated structure factor amplitudes, respectively) for data between 6.0- and 2.2-angstrom resolution for which F greater-than-or-equal-to 2-sigma(F). This is an antiparallel beta-barrel structure that incorporates "Greek key" and "jellyroll" topological motifs. A shallow depression is formed at the active site by part of the beta-sheet and an OMEGA-loop flanking one side of the sheet. His83, the histidyl residue which is the phosphorylation target of HPr and which transfers the phosphoryl group to the IIB domain of the permease, is located at the C-terminus of a beta-strand. The N-epsilon atom is partially solvated and also interacts with the N-epsilon atom of a second histidyl residue, His68, located at the N-terminus of an adjacent beta-strand, suggesting they share a proton. The geometry of the hydrogen bond is imperfect, though. Electrostatic interactions with other polar groups and van der Waals contacts with the side chains of two flanking phenylalanine residues assure the precise orientation of the imidazole rings. The hydrophobic nature of the surface around the His83-His68 pair may be required for protein-protein recognition by HPr or/and by the IIB domain of the permease. The side chains of two aspartyl residues, Asp31 and Asp87, are oriented toward each other across a narrow groove, about 7 angstrom from the active-site His83, suggesting they may play a role in protein-protein interaction. A model of the phosphorylated form of the molecule is proposed, in which oxygen atoms of the phosphoryl group interact with the side chain of His68 and with the main-chain nitrogen atom of a neighboring residue, Val89. The model, in conjunction with previously reported site-directed mutagenesis experiments, suggests that the phosphorylation of His83 may be accompanied by the protonation of His68. This may be important for the interaction with the IIB domain of the permease and/or play a catalytic role in the phosphoryl transfer from IIA to IIB.