Mannose permease of Escherichia coli. Domain structure and function of the phosphorylating subunit.
The mannose permease of Escherichia coli is a component of the phosphotransferase system. It transports mannose and related hexoses by a mechanism that couples sugar transport with sugar phosphorylation. It is a complex consisting of two transmembrane subunits (II-PMan and II-MMan) and a hydrophilic subunit (IIIMan). IIIMan also exists in a soluble form as dimer in the cytoplasm. Each monomer of IIIMan consists of two structurally and functionally distinct domains which are linked by a flexible hinge of the sequence KAAPAPAAAAPKAAPTPAKP. Both domains are transiently phosphorylated. The NH2-terminal domain (P13) is phosphorylated at N-3 of His-10 by the cytoplasmic phosphorylcarrier protein phospho-HPr. The COOH-terminal domain (P20) is phosphorylated by P13 at N-1 of His-175. Phosphoryltransfer occurs not only between P13 and P20 on the same IIIMan subunit but also between isolated domains and between domains on different subunits of the dimer. In the presence of the IIMan subunits, the phosphoryl group is directly transferred from His-175 of P20 to the sugar substrates of the permease. The P13 domain contains the contact sites for dimerization of IIIMan. The P20 domain contains the contact sites for interaction with the IIMan subunits. By reconstructing the ptsL gene, the two domains were expressed as individual polypeptides and the length of the hinge between P13 and P20 was changed. The in vivo and in vitro activities of mutant IIIMan were little affected by these modifications. The hinge is highly sensitive to proteolytic cleavage in vitro and its specificity for proteases can be modified by introducing the appropriate specificity determinants.
Erni
B
B
Department of Microbiology, University of Basel, Switzerland.
Zanolari
B
B
Graff
P
P
Kocher
H P
HP
D000595:Amino Acid Sequence; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006639:Histidine / Q000378:metabolism; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D010446:Peptide Fragments / Q000378:metabolism; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D014357:Trypsin / Q000378:metabolism
D004269:DNA, Bacterial; D046911:Macromolecular Substances; D010446:Peptide Fragments; D006639:Histidine; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C037992:phosphoenolpyruvate-mannose phosphotransferase; D014357:Trypsin