Regulation of the maltose transport system of Escherichia coli by the glucose-specific enzyme III of the phosphoenolpyruvate-sugar phosphotransferase system. Characterization of inducer exclusion-resistant mutants and reconstitution of inducer exclusion in proteoliposomes.
Maltose transport in Escherichia coli is regulated at the protein level by the glucose-specific enzyme III (IIIglc) of the phosphoenolpyruvate-sugar phosphotransferase system, by a mechanism known as inducer exclusion. We have isolated and characterized four mutants in the maltose transport system, all of which are in malK, which are resistant to inducer exclusion. The mutations in three of these mutants fall within the COOH-terminal domain of MalK and suggest the first reported function for this domain. Two of these are in a region which shows sequence similarity to lacY and melB, both of which are also regulated by IIIglc, and thus may define a IIIglc-binding domain. We have also reconstituted inducer exclusion in proteoliposomes made from membranes overexpressing the maltose permease. Maltose transport is inhibited by 50-60% when IIIglc is included in the intravesicular space. The inhibition is due to a decrease in the Vmax of transport by a factor of 2. IIIglc does not affect the coupling of ATP hydrolysis to maltose transport, since the ratio of ATP hydrolyzed/maltose transported remained constant in the presence and absence of IIIglc. Finally, the Ki for IIIglc was 40 microM, roughly the same as the in vivo concentration of IIIglc.
Dean
D A
DA
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Reizer
J
J
Nikaido
H
H
Saier
M H
MH
D000595:Amino Acid Sequence; D001483:Base Sequence; D001692:Biological Transport; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D005838:Genotype; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism; D008320:Maltose / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000235:genetics / Q000378:metabolism*; D011510:Proteolipids / Q000378:metabolism
D029968:Escherichia coli Proteins; D008081:Liposomes; D015345:Oligonucleotide Probes; D011510:Proteolipids; C032242:crr protein, E coli; C020485:proteoliposomes; D008320:Maltose; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System