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High-throughput assays for SAM-dependent methyltransferases: advances, challenges, and future perspectives
Renia Fotiadou1, Florian Heinz2, Konstantin F G Weigmann2
1Department of Chemistry, University of Crete, Voutes University Campus, 70013 Heraklion, Greece. ipavlidis@uoc.gr.
Natural Product Reports
|May 5, 2026
Summary
Methyltransferases are diverse enzymes that methylate substrates using S-adenosyl-L-methionine. This review examines high-throughput screening assays for monitoring their activity, crucial for enzyme engineering.
Area of Science:
- Biochemistry and enzymology
- Molecular biology
- Enzyme kinetics
Background:
- Methyltransferases (EC 2.1.1.-) are functionally diverse enzymes found across all life domains, catalyzing essential methylation reactions.
- Despite substrate diversity, most methyltransferases use S-adenosyl-L-methionine (SAM) as a common methyl donor.
- Existing assays often struggle with complex substrates and regiospecificity, limiting enzyme engineering applications.
Purpose of the Study:
- To critically review high-throughput screening assays for methyltransferase activity.
- To address the need for efficient analysis in enzyme engineering.
- To cover advancements in methyltransferase assay development up to January 2026.
Main Methods:
- Review of colorimetric and fluorometric assays for methyltransferase activity.
- Analysis of strategies based on detecting S-adenosyl-L-homocysteine (SAH) or its degradation products.
- Evaluation of assays for purified enzymes versus crude cell lysates.
Main Results:
- Colorimetric and fluorometric assays are widely used but have limitations.
- Detection of SAH or its by-products is a common strategy.
- Sophisticated tools are needed for regiospecificity determination in complex methylations.
Conclusions:
- High-throughput screening assays are essential for exploiting methyltransferases, especially in enzyme engineering.
- Continued development of robust and specific assays is needed.
- This review provides a critical overview of current assay technologies.
