Membrane localization itself but not binding to IICB is directly responsible for the inactivation of the global repressor Mlc in Escherichia coli.
Mlc is a global transcriptional repressor involved in the regulation of genes linked to glucose metabolism. The activity of Mlc is modulated through the interaction with a major glucose transporter, IICBGlc, in response to external glucose. To understand how IICBGlc-Mlc interaction controls the repressor activity of Mlc, we attempted to isolate Mlc mutants that retain the ability to repress target genes even in the presence of glucose. The Mlc mutants were tested for their ability to interact with IICBGlc. Mutants in which a single amino acid substitution occurs in the N-terminal portion were no longer able to bind to IICBGlc, suggesting that the N-terminal region of Mlc is primarily responsible for the interaction with IICBGlc. To examine whether the Mlc-IICBGlc interaction and/or the membrane localization of Mlc per se are essential for the inactivation of Mlc, the properties of several hybrid proteins in which either IIBGlc or Mlc is fused to membrane proteins were analysed. The cytoplasmic IIBGlc domain failed to inhibit the Mlc action although it retains the ability to bind Mlc in cells. However, it gained the ability to inhibit the Mlc activity when it was fused to a membrane protein LacY. In addition, we showed that Mlc is inactivated when fused to membrane proteins but not when fused to cytoplasmic proteins. We conclude that the IICBGlc-Mlc interaction is dispensable for the inactivation of Mlc, and that membrane localization is directly responsible for the inactivation of Mlc.
Tanaka
Yuya
Y
Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa, Nagoya 464-8602, Japan.
Itoh
Fumie
F
Kimata
Keiko
K
Aiba
Hiroji
H
D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000037:antagonists & inhibitors*; D005838:Genotype; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism*; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012329:RNA, Bacterial / Q000235:genetics; D012097:Repressor Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014158:Transcription, Genetic
D017931:DNA Primers; D029968:Escherichia coli Proteins; C111065:Mlc protein, E coli; D009004:Monosaccharide Transport Proteins; D012329:RNA, Bacterial; D012097:Repressor Proteins; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C031629:phosphoenolpyruvate-glucose phosphotransferase