Limits...
Autoregulation of the Escherichia coli melR promoter: repression involves four molecules of MelR.

Samarasinghe S, El-Robh MS, Grainger DC, Zhang W, Soultanas P, Busby SJ - Nucleic Acids Res. (2008)

Bottom Line: Optimal repression requires MelR binding to a site that overlaps the melR transcription start point and to upstream sites.In this work, we have investigated the different determinants needed for optimal repression and their spatial requirements.We show that repression requires a complex involving four DNA-bound MelR molecules, and that the global CRP regulator plays little or no role.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, The University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.

ABSTRACT
The Escherichia coli MelR protein is a transcription activator that autoregulates its own promoter by repressing transcription initiation. Optimal repression requires MelR binding to a site that overlaps the melR transcription start point and to upstream sites. In this work, we have investigated the different determinants needed for optimal repression and their spatial requirements. We show that repression requires a complex involving four DNA-bound MelR molecules, and that the global CRP regulator plays little or no role.

Show MeSH

Related in: MedlinePlus

Binding of MelR to the TB22 and TB28 promoter fragments. The figure shows autoradiograms of electromobility shift assays performed with labelled TB22 and TB28 fragments, as indicated, incubated with no MelR (lane 1) or 2000 nM (lane 2), 1000 nM (lane 3), 500 nM (lane 4), 250 nM (lane 5), 125 nM (lane 6), 62.5 nM (lane 7), 31.25 nM (lane 8) and 15.62 nM (lane 9) purified MelR.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2377442&req=5

Figure 7: Binding of MelR to the TB22 and TB28 promoter fragments. The figure shows autoradiograms of electromobility shift assays performed with labelled TB22 and TB28 fragments, as indicated, incubated with no MelR (lane 1) or 2000 nM (lane 2), 1000 nM (lane 3), 500 nM (lane 4), 250 nM (lane 5), 125 nM (lane 6), 62.5 nM (lane 7), 31.25 nM (lane 8) and 15.62 nM (lane 9) purified MelR.

Mentions: Our results show that MelR-dependent repression of the melR promoter is contingent on MelR binding to site R and is somehow modulated by MelR binding to upstream sites 2, 1 and 1′. Hence an electromobility shift assay (Figure 7) and a DNAase I footprinting experiment (Figure 8) were used to investigate the occupation of the different binding sites at the TB22 and TB28 promoters by purified MelR.Figure 7.


Autoregulation of the Escherichia coli melR promoter: repression involves four molecules of MelR.

Samarasinghe S, El-Robh MS, Grainger DC, Zhang W, Soultanas P, Busby SJ - Nucleic Acids Res. (2008)

Binding of MelR to the TB22 and TB28 promoter fragments. The figure shows autoradiograms of electromobility shift assays performed with labelled TB22 and TB28 fragments, as indicated, incubated with no MelR (lane 1) or 2000 nM (lane 2), 1000 nM (lane 3), 500 nM (lane 4), 250 nM (lane 5), 125 nM (lane 6), 62.5 nM (lane 7), 31.25 nM (lane 8) and 15.62 nM (lane 9) purified MelR.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2377442&req=5

Figure 7: Binding of MelR to the TB22 and TB28 promoter fragments. The figure shows autoradiograms of electromobility shift assays performed with labelled TB22 and TB28 fragments, as indicated, incubated with no MelR (lane 1) or 2000 nM (lane 2), 1000 nM (lane 3), 500 nM (lane 4), 250 nM (lane 5), 125 nM (lane 6), 62.5 nM (lane 7), 31.25 nM (lane 8) and 15.62 nM (lane 9) purified MelR.
Mentions: Our results show that MelR-dependent repression of the melR promoter is contingent on MelR binding to site R and is somehow modulated by MelR binding to upstream sites 2, 1 and 1′. Hence an electromobility shift assay (Figure 7) and a DNAase I footprinting experiment (Figure 8) were used to investigate the occupation of the different binding sites at the TB22 and TB28 promoters by purified MelR.Figure 7.

Bottom Line: Optimal repression requires MelR binding to a site that overlaps the melR transcription start point and to upstream sites.In this work, we have investigated the different determinants needed for optimal repression and their spatial requirements.We show that repression requires a complex involving four DNA-bound MelR molecules, and that the global CRP regulator plays little or no role.

View Article: PubMed Central - PubMed

Affiliation: School of Biosciences, The University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.

ABSTRACT
The Escherichia coli MelR protein is a transcription activator that autoregulates its own promoter by repressing transcription initiation. Optimal repression requires MelR binding to a site that overlaps the melR transcription start point and to upstream sites. In this work, we have investigated the different determinants needed for optimal repression and their spatial requirements. We show that repression requires a complex involving four DNA-bound MelR molecules, and that the global CRP regulator plays little or no role.

Show MeSH
Related in: MedlinePlus