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Recognition of dual symmetry by the controller protein C.Esp1396I based on the structure of the transcriptional activation complex.

McGeehan JE, Ball NJ, Streeter SD, Thresh SJ, Kneale GG - Nucleic Acids Res. (2011)

Bottom Line: The molecular recognition of promoter sequences by such transcriptional regulators is poorly understood, in part because the DNA sequence motifs do not conform to a well-defined symmetry.The structure reveals how two different symmetries within the operator are simultaneously recognized by the homo-dimeric protein, underpinned by a conformational change in one of the protein subunits.The recognition of two different DNA symmetries through movement of a flexible loop in one of the protein subunits may represent a general mechanism for the recognition of pseudo-symmetric DNA sequences.

View Article: PubMed Central - PubMed

Affiliation: Biomolecular Structure Group, Institute of Biomedical and Biomolecular Sciences, School of Biological Sciences, University of Portsmouth, Portsmouth, Hampshire PO1 2DY, UK.

ABSTRACT
The controller protein C.Esp1396I regulates the timing of gene expression of the restriction-modification (RM) genes of the RM system Esp1396I. The molecular recognition of promoter sequences by such transcriptional regulators is poorly understood, in part because the DNA sequence motifs do not conform to a well-defined symmetry. We report here the crystal structure of the controller protein bound to a DNA operator site. The structure reveals how two different symmetries within the operator are simultaneously recognized by the homo-dimeric protein, underpinned by a conformational change in one of the protein subunits. The recognition of two different DNA symmetries through movement of a flexible loop in one of the protein subunits may represent a general mechanism for the recognition of pseudo-symmetric DNA sequences.

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The conserved DNA sequences obey different symmetries. (a) The C-boxes are symmetrical if the pseudo-dyad axis is placed between the central GT. (b) If the pseudo-dyad axis is placed at the central T, the C-boxes are no longer symmetrical but the other conserved element are. The pseudo dyad axes within operators (blue) and between operators (red) are shown as dotted lines. Figure reproduced from McGeehan et al. (2008). (c) C-proteins act as a genetic switch regulating the timing and expression of R–M genes. The −35 (green) and −10 (red) regions are indicated upstream of the C-gene (light blue) and the R-gene (data not shown). C-protein dimers are shown in blue and the sigma subunit of RNA polymerase in orange. C-protein is expressed at low levels from a weak C-independent promoter (data not shown). A C-protein dimer first occupies the high-affinity OL site and stimulates transcription of the C-gene through recruitment of RNA polymerase sigma subunit to the −35 site. As the C-protein concentration increases, a dimer occupies the OR site and occludes the −35 site down-regulating the expression of the C- and R-genes. Adapted from McGeehan et al. (11).
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gkr1250-F1: The conserved DNA sequences obey different symmetries. (a) The C-boxes are symmetrical if the pseudo-dyad axis is placed between the central GT. (b) If the pseudo-dyad axis is placed at the central T, the C-boxes are no longer symmetrical but the other conserved element are. The pseudo dyad axes within operators (blue) and between operators (red) are shown as dotted lines. Figure reproduced from McGeehan et al. (2008). (c) C-proteins act as a genetic switch regulating the timing and expression of R–M genes. The −35 (green) and −10 (red) regions are indicated upstream of the C-gene (light blue) and the R-gene (data not shown). C-protein dimers are shown in blue and the sigma subunit of RNA polymerase in orange. C-protein is expressed at low levels from a weak C-independent promoter (data not shown). A C-protein dimer first occupies the high-affinity OL site and stimulates transcription of the C-gene through recruitment of RNA polymerase sigma subunit to the −35 site. As the C-protein concentration increases, a dimer occupies the OR site and occludes the −35 site down-regulating the expression of the C- and R-genes. Adapted from McGeehan et al. (11).

Mentions: Earlier biochemical and biophysical analysis in our laboratory suggested the basis of the genetic switch in AhdI (11–15). Low-level expression of the C-protein from a weak promoter leads to a delay in transcription until sufficient protein accumulates to form a functional dimer. The C-protein dimer activates transcription of the C/R operon, forming a positive feedback loop, which leads to a rapid increase in C-protein expression; at higher concentrations, a second dimer is recruited to the promoter, displacing the σ subunit of RNA polymerase and thereby repressing transcription of its own gene (and hence expression of the R gene) in a negative feedback loop (Figure 1). A similar, but more complex, mechanism has been proposed for the R–M system Esp1396I. Experiments conducted in collaboration with Severinov and colleagues (16) have shown that in the R–M system Esp1396I, the C-protein, in addition to regulation of the R gene, represses the M gene by binding as a dimer to a high-affinity site that overlaps the transcriptional start site.Figure 1.


Recognition of dual symmetry by the controller protein C.Esp1396I based on the structure of the transcriptional activation complex.

McGeehan JE, Ball NJ, Streeter SD, Thresh SJ, Kneale GG - Nucleic Acids Res. (2011)

The conserved DNA sequences obey different symmetries. (a) The C-boxes are symmetrical if the pseudo-dyad axis is placed between the central GT. (b) If the pseudo-dyad axis is placed at the central T, the C-boxes are no longer symmetrical but the other conserved element are. The pseudo dyad axes within operators (blue) and between operators (red) are shown as dotted lines. Figure reproduced from McGeehan et al. (2008). (c) C-proteins act as a genetic switch regulating the timing and expression of R–M genes. The −35 (green) and −10 (red) regions are indicated upstream of the C-gene (light blue) and the R-gene (data not shown). C-protein dimers are shown in blue and the sigma subunit of RNA polymerase in orange. C-protein is expressed at low levels from a weak C-independent promoter (data not shown). A C-protein dimer first occupies the high-affinity OL site and stimulates transcription of the C-gene through recruitment of RNA polymerase sigma subunit to the −35 site. As the C-protein concentration increases, a dimer occupies the OR site and occludes the −35 site down-regulating the expression of the C- and R-genes. Adapted from McGeehan et al. (11).
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Related In: Results  -  Collection

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gkr1250-F1: The conserved DNA sequences obey different symmetries. (a) The C-boxes are symmetrical if the pseudo-dyad axis is placed between the central GT. (b) If the pseudo-dyad axis is placed at the central T, the C-boxes are no longer symmetrical but the other conserved element are. The pseudo dyad axes within operators (blue) and between operators (red) are shown as dotted lines. Figure reproduced from McGeehan et al. (2008). (c) C-proteins act as a genetic switch regulating the timing and expression of R–M genes. The −35 (green) and −10 (red) regions are indicated upstream of the C-gene (light blue) and the R-gene (data not shown). C-protein dimers are shown in blue and the sigma subunit of RNA polymerase in orange. C-protein is expressed at low levels from a weak C-independent promoter (data not shown). A C-protein dimer first occupies the high-affinity OL site and stimulates transcription of the C-gene through recruitment of RNA polymerase sigma subunit to the −35 site. As the C-protein concentration increases, a dimer occupies the OR site and occludes the −35 site down-regulating the expression of the C- and R-genes. Adapted from McGeehan et al. (11).
Mentions: Earlier biochemical and biophysical analysis in our laboratory suggested the basis of the genetic switch in AhdI (11–15). Low-level expression of the C-protein from a weak promoter leads to a delay in transcription until sufficient protein accumulates to form a functional dimer. The C-protein dimer activates transcription of the C/R operon, forming a positive feedback loop, which leads to a rapid increase in C-protein expression; at higher concentrations, a second dimer is recruited to the promoter, displacing the σ subunit of RNA polymerase and thereby repressing transcription of its own gene (and hence expression of the R gene) in a negative feedback loop (Figure 1). A similar, but more complex, mechanism has been proposed for the R–M system Esp1396I. Experiments conducted in collaboration with Severinov and colleagues (16) have shown that in the R–M system Esp1396I, the C-protein, in addition to regulation of the R gene, represses the M gene by binding as a dimer to a high-affinity site that overlaps the transcriptional start site.Figure 1.

Bottom Line: The molecular recognition of promoter sequences by such transcriptional regulators is poorly understood, in part because the DNA sequence motifs do not conform to a well-defined symmetry.The structure reveals how two different symmetries within the operator are simultaneously recognized by the homo-dimeric protein, underpinned by a conformational change in one of the protein subunits.The recognition of two different DNA symmetries through movement of a flexible loop in one of the protein subunits may represent a general mechanism for the recognition of pseudo-symmetric DNA sequences.

View Article: PubMed Central - PubMed

Affiliation: Biomolecular Structure Group, Institute of Biomedical and Biomolecular Sciences, School of Biological Sciences, University of Portsmouth, Portsmouth, Hampshire PO1 2DY, UK.

ABSTRACT
The controller protein C.Esp1396I regulates the timing of gene expression of the restriction-modification (RM) genes of the RM system Esp1396I. The molecular recognition of promoter sequences by such transcriptional regulators is poorly understood, in part because the DNA sequence motifs do not conform to a well-defined symmetry. We report here the crystal structure of the controller protein bound to a DNA operator site. The structure reveals how two different symmetries within the operator are simultaneously recognized by the homo-dimeric protein, underpinned by a conformational change in one of the protein subunits. The recognition of two different DNA symmetries through movement of a flexible loop in one of the protein subunits may represent a general mechanism for the recognition of pseudo-symmetric DNA sequences.

Show MeSH