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Transmembrane Domain Oligomerization Propensity determined by ToxR Assay
Published on: May 26, 2011
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An oligomeric switch controls the Mrr-induced SOS response in E. coli
Anaïs C Bourges1, Oscar E Torres Montaguth2, Wubishet Tadesse2
1Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy, NY, 12180, USA; Centre de Biochimie Structurale, CNRS, INSERM, Université de Montpellier, 34090, Montpellier, France.
DNA Repair
|November 21, 2020
Summary
Mrr endonuclease, an Escherichia coli protein, shifts from inactive tetramers to active dimers under stress. This oligomeric switch, triggered by specific enzymes or high pressure, reveals insights into DNA damage and host response mechanisms.
Area of Science:
- Molecular Biology
- Biochemistry
- Genetics
Background:
- Mrr endonuclease from Escherichia coli K12 is a type IV restriction enzyme.
- It cleaves foreign methylated DNA but can also damage host chromosomal DNA, inducing the SOS response.
- Mrr activation involves an oligomeric switch from inactive tetramers to active dimers.
Purpose of the Study:
- To investigate the in vivo stoichiometry of Mrr mutants using scanning number and brightness (sN&B) analysis.
- To examine Mrr mutants' response to M.HhaII activity and high pressure (HP) shock.
- To elucidate the relationship between Mrr subunit interactions, activity, and DNA binding.
Main Methods:
- Scanning number and brightness (sN&B) analysis to determine in vivo protein stoichiometry.
- High-pressure fluorescence correlation spectroscopy (HP FCS) to observe in vitro tetramer dissociation.
- Structural modeling of the Mrr tetramer bound to DNA.
Main Results:
- sN&B analysis determined the in vivo stoichiometry of a constitutively active Mrr mutant.
- HP FCS demonstrated direct pressure-induced dissociation of purified GFP-Mrr tetramers.
- A structural model revealed DNA-bound tetramer sequestration and dimer activation.
Conclusions:
- Mrr's oligomeric switch from tetramer to dimer is crucial for its activity.
- Specific methyltransferases and high pressure trigger this switch, leading to DNA damage.
- Structural insights explain how DNA binding and subunit dissociation regulate Mrr endonuclease activity.
Keywords:
Live cell imagingMethyl-directed endonucleasePressure-induced SOS-responseProtein modelingQuantitative fluorescence fluctuation microscopy and spectroscopyMore Related Videos
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