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Updated: Jul 22, 2026
11:42
Iterative Optimization of DNA Duplexes for Crystallization of SeqA-DNA Complexes
Published on: November 1, 2012
Substituting an alpha-helix switches the sequence-specific DNA interactions of a repressor
Cell
|September 1, 1984
Summary
Researchers identified DNA-recognition helices in bacteriophage 434 repressor and cro proteins. A hybrid protein showed that swapping these helices altered DNA binding specificity, confirming their role.
Area of Science:
- Molecular Biology
- Genetics
- Protein-DNA Interactions
Background:
- Many DNA-binding proteins utilize alpha-helices for specific DNA sequence recognition.
- The repressor and cro proteins of bacteriophage 434 are structurally related but exhibit distinct DNA-binding properties.
- These proteins interact with multiple sites within the phage 434 operator (OR).
Purpose of the Study:
- To identify the presumptive DNA-recognition alpha-helices in bacteriophage 434 repressor and cro proteins using amino acid sequence homology.
- To investigate the role of these putative helices in determining DNA-binding specificity.
- To engineer a hybrid protein to test the function of the identified helices.
Main Methods:
- Amino acid sequence homology analysis to predict DNA-recognition helices.
- Chemical probing (dimethyl sulfate) to map protein-DNA contacts.
- Protein engineering to create a hybrid repressor* by swapping putative recognition helices.
Main Results:
- The study identified presumptive DNA-recognition alpha-helices in both 434 repressor and cro proteins.
- Dimethyl sulfate probing revealed distinct DNA-binding contacts for repressor and cro proteins at operator sites.
- The engineered hybrid protein, repressor*, exhibited DNA-binding contacts characteristic of the cro protein, not the original repressor.
Conclusions:
- The identified alpha-helices are crucial for determining the specific DNA sequence contacts made by bacteriophage 434 repressor and cro proteins.
- Swapping these helices effectively transfers the DNA-binding specificity from one protein to another.
- This study provides direct evidence for the role of specific alpha-helices in sequence-selective DNA recognition by these viral proteins.
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