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Cloning and structure of the BepI modification methylase
D Kupper1, J G Zhou, P Venetianer
1Biological Research Center, Hungarian Academy of Sciences, Szeged.
Nucleic Acids Research
|February 11, 1989
Summary
The Brevibacterium epidermidis BepI methylase gene was cloned and expressed in E. coli. This cytosine-specific DNA methylase, BepI, recognizes the CGCG sequence and methylates the external cytosine.
Area of Science:
- Molecular Biology
- Enzymology
- Genetics
Background:
- Brevibacterium epidermidis harbors restriction-modification systems.
- The FnuDII isoschizomer BepI endonuclease was previously isolated from this strain.
- The cognate methylase for BepI endonuclease was yet to be identified.
Purpose of the Study:
- To clone and characterize the gene encoding the CGCG-specific DNA methylase from Brevibacterium epidermidis.
- To investigate the expression and properties of the BepI methylase.
- To determine the nucleotide sequence and analyze the amino acid sequence of the BepI methylase.
Main Methods:
- Gene cloning in E. coli.
- Nucleotide sequencing.
- Bioinformatic analysis of amino acid sequence.
- Enzyme activity assays (implied).
Main Results:
- The gene for BepI methylase was successfully cloned and expressed in E. coli.
- Expression was dependent on the orientation of the cloned fragment.
- The BepI methylase gene predicts a protein of 403 amino acids.
- Sequence analysis revealed similarities to other cytosine methylases.
- BepI methylase was confirmed to methylate the external cytosine in its CGCG recognition sequence.
Conclusions:
- The BepI methylase gene from Brevibacterium epidermidis has been cloned and characterized.
- BepI methylase is a cytosine-specific DNA methylase likely cognate to BepI endonuclease.
- The enzyme's sequence shares homology with other methyltransferases, indicating conserved functional domains.