Antagonistic effects of dual PTS-catalysed phosphorylation on the Bacillus subtilis transcriptional activator LevR.
LevR, which controls the expression of the levoperon of Bacillus subtilis, is a regulatory protein containing an N-terminal domain similar to the NifA/NtrC transcriptional activator family and a C-terminal domain similar to the regulatory part of bacterial anti-terminators, such as BgIG and LicT. Here, we demonstrate that the activity of LevR is regulated by two phosphoenolpyruvate (PEP)-dependent phosphorylation reactions catalysed by the phosphotransferase system (PTS), a transport system for sugars, polyols and other sugar derivatives. The two general components of the PTS, enzyme I and HPr, and the two soluble, sugar-specific proteins of the lev-PTS, LevD and LevE, form a signal transduction chain allowing the PEP-dependent phosphorylation of LevR, presumably at His-869. This phosphorylation seems to inhibit LevR activity and probably regulates the induction of the lev operon. Mutants in which His-869 of LevR has been replaced with a non-phosphorylatable alanine residue exhibited constitutive expression from the lev promoter, as do levD or levE mutants. In contrast, PEP-dependent phosphorylation of LevR in the presence of only the general components of the PTS, enzyme I and HPr, regulates LevR activity positively. This phosphorylation most probably occurs at His-585. Mutants in which His-585 has been replaced with an alanine had lost stimulation of LevR activity and PEP-dependent phosphorylation by enzyme I and HPr. This second phosphorylation of LevR at His-585 is presumed to play a role in carbon catabolite repression.
Martin-Verstraete
I
I
Unité de Biochimie Microbienne, Institut Pasteur, URA 1300 du CNRS, Paris, France. iverstra@pasteur.fr
Charrier
V
V
Stülke
J
J
Galinier
A
A
Erni
B
B
Rapoport
G
G
Deutscher
J
J
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D010770:Phosphotransferases / Q000032:analysis / Q000235:genetics*; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
D001426:Bacterial Proteins; D014157:Transcription Factors; C067962:LevR protein, Bacillus subtilis; D010770:Phosphotransferases