1.5Å STRUCTURE OF THE ASP46 HPr MUTANT FROM ESCHERICHIA COLI. Scott Napper, Bruce Waygood, J. Wilson Quail, Louis T. J., Delbaere, Departments of Biochemistry and Chemistry, University of Saskatchewan, Saskatoon, Saskatchewan

Regulation of the phosphoenolpyruvate:sugar transferase system (PTS) in bacteria differs in Gram-positive to Gram-negative species. Gram-positive bacteria possess a regulatory mechanism at the level of HPr which is not seen in Gram-negative bacteria. Gram-positive sugar transport function can be inhibited through reversible phosphorylation of a conserved Ser46 residue of HPr. The kinase catalyzing this reaction is absent from Gram-negative bacteria and E. coli HPr is unable to be phosphorylated by it in vitro. The Asp46 mutant was created in attempt to mimic this regulatory phosphorylation event through similar introduction of negative charge. The mutant shows very similar properties to the phosphorylated Ser46 HPr in diminishing phosphotransfer activity. The crystallographic structure of this mutant has been determined through the method of molecular replacement and refined to a conventional R-index of 18.9%. The crystal structure shows that this inhibition occurs in the absence of any structural alterations. Rather it appears as though changes in the electrostatic surface potential are responsible for the inability of the protein to interact with other proteins, in particular Enzyme I.