The Bacillus stearothermophilus mannitol regulator, MtlR, of the phosphotransferase system. A DNA-binding protein, regulated by HPr and iicbmtl-dependent phosphorylation.
D-Mannitol is taken up by Bacillus stearothermophilus and phosphorylated via a phosphoenolpyruvate-dependent phosphotransferase system (PTS). The genes involved in the mannitol uptake were recently cloned and sequenced. One of the genes codes for a putative transcriptional regulator, MtlR. The presence of a DNA binding helix-turn-helix motif and two antiterminator-like PTS regulation domains, suggest that MtlR is a DNA-binding protein, the activity of which can be regulated by phosphorylation by components of the PTS. To demonstrate DNA binding of MtlR to a region upstream of the mannitol promoter, by DNA footprinting, MtlR was overproduced and purified. EI, HPr, IIAmtl, and IICBmtl of B. stearothermophilus were purified and used to demonstrate that MtlR can be phosphorylated and regulated by HPr and IICBmtl, in vitro. Phosphorylation of MtlR by HPr increases the affinity of MtlR for its binding site, whereas phosphorylation by IICBmtl results in a reduction of this affinity. The differential effect of phosphorylation, by two different proteins, on the DNA binding properties of a bacterial transcriptional regulator has not, to our knowledge, been described before. Regulation of MtlR by two components of the PTS is an example of an elegant control system sensing both the presence of mannitol and the need to utilize this substrate.
Henstra
S A
SA
Departments of Biochemistry, the Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Tuinhof
M
M
Duurkens
R H
RH
Robillard
G T
GT
D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D029968:Escherichia coli Proteins; D001411:Geobacillus stearothermophilus / Q000235:genetics / Q000378:metabolism*; D008353:Mannitol / Q000378:metabolism*; D009004:Monosaccharide Transport Proteins; D009876:Operon; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D012097:Repressor Proteins / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic