Mutational analysis of the enzyme III Glc of the phosphoenolpyruvate phosphotransferase system in Escherichia coli

Abstract

The phosphoenolpyruvate phosphotransferase system (PTS) component EIII Glc is responsible for transport and phosphorylation of glucose via EII Glc. It also regulates the catabolism of other carbon sources, such as lactose and maltose, by modulating both the intracellular concentrations of the corresponding inducers and of cAMP. Mutational analysis of EIII Glc was performed in order to identify crucial residues mediating the interactions between EIII Glc and its target proteins. Such mutations were isolated by in vitro hydroxylamine mutagenesis of the cloned EIII Glc gene, crr. Five mutated EIII Glc impaired in the function of inducer exclusion were obtained. However, these mutations did not abolish the function of EIII Glc in the transport and phosphorylation of glucose, nor in activation of adenylate cyclase. A single amino acid change was found for each mutation, which is located in a restricted area of the polypeptide chain: Gly 47 → Ser 47 for the HA2 and HA5 mutations, Ala 76 → Thr 76 for HA4 mutation and Ser 78 → Phe 78 for HA3 mutation, indicative of quaternary interactions between the corresponding region of EIII Glc and its target protein(s).