A nonradioactive DNA methyltransferase assay adaptable to high-throughput screening |
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Authors: | Woo Youn-Hi Rajagopalan P T Ravi Benkovic Stephen J |
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Affiliation: | 414 Wartik Laboratory, Department of Chemistry, Pennsylvania State University, University Park, PA 16802, USA. |
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Abstract: | We have developed a nonradioactive assay method for DNA methyltransferases based on the ability to protect substrate DNA from restriction. DNA immobilized to a microplate well was treated sequentially with methyltransferase and an appropriate endonuclease. The amount of methylated DNA product is reflected by a proportional decrease in endonuclease cleavage, which is in turn reflected by increased retention of the end-labeled affinity probe. A single universal substrate was designed to assay multiple methyltransferases including those that do not have a cognate endonuclease. The methodology developed is suited to screen a large number of compounds for inhibitors of various methyltransferases. |
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Keywords: | DNA methyltransferase Restriction/modification system DNA immobilization |
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