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Interaction with DNA of oligopeptides related to the Arc repressor
N Helbecque1, A el Idrissi Boutaher, J P Hénichart
1Service d'Epidémiologie et de Santé Publique et INSERM, Institut Pasteur de Lille, France.
Abstract:
Three different peptides, A13, A14x2 and A20, related to the Arc repressor from Salmonella phage P22, were synthesized. They all contained the 13 N-terminal residues of Arc known to form a beta-sheet interacting with the operator OArc. In the case of A20, the tripeptide Lys-Trp-Lys was added in the C-terminal position because of its propensity to increase affinity to DNA. The interaction of the three peptides with OArc and with other related (OMnt from the same phage) and unrelated oligonucleotides was followed using circular dichroism, filter binding assays and DNaseI protection experiments. While Kd = 10(-9) M for the protein, 8.7 x 10(-5) M and 7.7 x 10(-6) M Kd values were obtained for A14x2 and A20 interacting with OArc.
Insights
Synthesized peptides mimicking the Arc repressor
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- The Arc repressor regulates gene expression in Salmonella phage P22.
- The N-terminal region of Arc forms a beta-sheet crucial for operator binding.
- Understanding peptide-DNA interactions is key to deciphering gene regulation mechanisms.
Purpose of the Study:
- To synthesize and characterize peptides based on the Arc repressor's N-terminal DNA-binding domain.
- To investigate the DNA-binding affinities of these synthetic peptides to the OArc operator sequence.
- To explore the role of specific amino acid modifications in peptide-DNA interactions.
Main Methods:
- Peptide synthesis of A13, A14x2, and A20.
- Circular dichroism spectroscopy to monitor structural changes.
- Filter binding assays and DNaseI protection experiments to quantify DNA binding.
Main Results:
- Peptides A14x2 and A20 showed measurable binding to the OArc operator.
- Dissociation constants (Kd) for A14x2 and A20 were determined as 8.7 x 10(-5) M and 7.7 x 10(-6) M, respectively.
- Peptide A20, with added Lys-Trp-Lys, exhibited enhanced DNA affinity compared to A14x2.
Conclusions:
- Synthetic peptides representing the Arc repressor's N-terminal beta-sheet can interact with OArc DNA.
- The binding affinity of these peptides is significantly lower than the native Arc repressor protein.
- Specific C-terminal modifications, like those in A20, can modulate peptide-DNA binding characteristics.