title: Exchange of phosphoryl groups between HPr molecules of the phosphoenolpyruvate-dependent phosphotransferase system is an autocatalytic process.
abstract: HPr, a central component of the phosphoenolpyruvate-dependent phosphotransferase system, can exist in Escherichia coli in a phosphorylated (PHPr) and a nonphosphorylated form. We show that, beside the normal transfer of the phosphoryl group from PHPr to enzymes II and III, PHPr can phosphorylate other HPr molecules in an autocatalytic exchange reaction. The reaction is very fast but is inhibited by labeling the protein with Bolton-Hunter reagent. We demonstrate that the exchange reaction can be used to determine the delta G degree of the phosphoryl group of mutant forms of PHPr relative to wild-type PHPr. Two HPr mutants were constructed by site-directed mutagenesis, HPr P11E and HPr E68A. Both show altered phosphoryl group potentials but show no significantly altered KM or Vmax values compared to wild-type HPr, illustrating the sensitivity of the exchange process. The exchange reaction does not occur between HPr from E. coli and HPr from Staphylococcus carnosus.
authors.affiliation: Department of Physical Chemistry, State University of Groningen, The Netherlands.
authors.forename: A A
authors.initials: AA
authors.lastname: van Dijk
authors.forename: R
authors.initials: R
authors.lastname: Eisermann
authors.forename: W
authors.initials: W
authors.lastname: Hengstenberg
authors.forename: G T
authors.initials: GT
authors.lastname: Robillard
mesh_terms: D001345:Autoradiography; D001426:Bacterial Proteins; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D013210:Staphylococcus / Q000201:enzymology; D014316:Tritium
chemical_list: D001426:Bacterial Proteins; D015345:Oligonucleotide Probes; D014316:Tritium; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr