This recombinant Gardnerella vaginalis 5-1 site-specific DNA-methyltransferase (adenine-specific) is expressed in a yeast system and contains the complete protein spanning amino acids 1-525. The protein includes an N-terminal 6xHis-tag for easier purification and detection. With purity levels exceeding 90% as confirmed by SDS-PAGE, this preparation appears well-suited for research applications.
DNA-methyltransferases catalyze the transfer of methyl groups to DNA, usually targeting adenine or cytosine bases. This process may influence gene expression and regulation in ways we're still learning to understand. Since this particular enzyme specifically methylates adenine residues, it could prove useful for investigating epigenetic modifications and how they might affect cellular processes. Research into these mechanisms seems increasingly important for advancing our knowledge of gene regulation and epigenetics.
Potential Applications
Note: The applications listed below are based on what we know about this protein's biological functions, published research, and experience from experts in the field. However, we haven't fully tested all of these applications ourselves yet. We'd recommend running some preliminary tests first to make sure they work for your specific research goals.
1. In Vitro DNA Methylation Assays
Researchers can use this recombinant adenine-specific DNA methyltransferase to examine site-specific DNA methylation patterns and kinetics under controlled laboratory conditions. Testing various DNA sequences and oligonucleotides might reveal the enzyme's substrate preferences and help identify favored recognition sites. The N-terminal 6xHis tag should make purification and immobilization for enzymatic assays more straightforward. Studies like these could shed light on bacterial DNA modification systems and their potential role in gene regulation.
2. Protein-DNA Interaction Studies
The 6xHis-tagged methyltransferase works well in electrophoretic mobility shift assays (EMSA) and surface plasmon resonance experiments when characterizing DNA-binding properties. Scientists can examine binding affinity, specificity, and kinetics using different DNA substrates and cofactors, though results may vary depending on experimental conditions. These investigations might help clarify the molecular mechanisms behind site recognition and binding for this bacterial methyltransferase. The high purity (>90%) likely contributes to more reliable and reproducible binding data.
3. Comparative Enzymology Research
This Gardnerella vaginalis methyltransferase could serve as a reference point for comparing other bacterial DNA methyltransferases. By analyzing structural and functional differences between methyltransferases from various bacterial species, researchers might gain insights into evolutionary relationships and functional diversification—though such comparisons can be complex. The full-length recombinant protein (1-525aa) provides both catalytic and regulatory domains, which appears necessary for thorough comparative analysis.
4. Antibody Development and Validation
The purified recombinant protein can function as an antigen for generating specific antibodies against Gardnerella vaginalis DNA methyltransferase. The 6xHis tag allows for straightforward immobilization on nickel-based surfaces during ELISA development and antibody screening assays. These antibodies would likely become valuable research tools for studying methyltransferase expression and localization in bacterial cultures, and they might help develop detection methods for microbiological research.
5. Biochemical Characterization Studies
This recombinant enzyme enables researchers to determine optimal reaction conditions, cofactor requirements, and kinetic parameters for the methylation reaction. Investigations can include pH and temperature optimization, metal ion dependencies, and inhibitor screening to characterize the enzyme's biochemical properties more completely. The yeast expression system may provide proper eukaryotic folding, potentially producing active enzyme suitable for detailed kinetic analysis and mechanism studies—though activity levels can sometimes vary between expression systems.
