MAPPING OF THE BINDING INTERFACES OF THE PROTEINS OF THE BACTERIAL PHOSPHOTRANSFERASE SYSTEM, HPR AND IIA(GLC)

Enzyme IIA(glc) and HPr are central regulatory and phosphocarrier proteins of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) of bacteria. During phosphoryl transfer from phosphoenolpyruvate to glucose, phosphate is transferred from HPr to enzyme IIA(glc). In order to characterize the binding interfaces of the two proteins during phosphate transfer, N-15-edited and N-15-filtered NMR experiments have been recorded for the complex of enzyme IIA(glc) and HPr from Bacillus subtilis. Uniformly N-15-labeled enzyme IIA(glc) and nonlabeled HPr were used in these studies. Residues which undergo significant chemical shift changes upon complex formation have been identified for both proteins. The binding interfaces of the two proteins, suggested by the observed chemical shift changes, involve predominantly hydrophobic surfaces near the active site His-15 of HPr and the phosphoryl acceptor His-83 of IIA(glc).