Operator-bound GalR dimers close DNA loops by direct interaction: tetramerization and inducer binding.
Szabolcs Semsey1, Mark Geanacopoulos, Dale E A Lewis
1Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute/ National Institutes of Health, Bethesda, MD 20892-4264, USA.
The EMBO Journal
|August 10, 2002
Summary
A GalR mutant protein in E. coli can repress gal operon transcription without the usual HU protein or supercoiled DNA. This suggests direct GalR dimer interaction drives DNA looping for gene repression.
Area of Science:
- Molecular Biology
- Microbiology
- Genetics
Background:
- The Gal repressosome complex in Escherichia coli regulates gal operon transcription.
- This complex involves DNA looping mediated by GalR dimers, the histone-like protein HU, and supercoiled DNA.
Purpose of the Study:
- To investigate the role of protein-protein interactions in Gal repressosome assembly.
- To characterize a GalR mutant with altered DNA-binding and repression capabilities.
Main Methods:
- Isolation and characterization of a specific GalR mutant (R282L).
- In vivo and in vitro transcription assays.
- DNA looping assays with and without HU and supercoiled DNA.
Main Results:
- The R282L GalR mutant repressed transcription independently of HU and supercoiled DNA.
- Repression by the mutant still required specific DNA operator sites (O(E) and O(I)).
- The mutation enhanced GalR dimer-dimer interaction, facilitating DNA looping.
Conclusions:
- GalR dimers directly interact to form the DNA loop, with HU and supercoiled DNA acting as stabilizers.
- The R282L mutation reveals a mechanism for direct protein-mediated DNA looping in gene regulation.
- Inducer binding likely modulates GalR tetramerization to control transcription.
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