Engineering and Directed Evolution of DNA Methyltransferases

  • Paola Laurino
  • Liat Rockah-Shmuel
  • Dan S. TawfikEmail author
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 945)


DNA methyltransferases (MTases) constitute an attractive target for protein engineering, thus opening the road to new ways of manipulating DNA in a unique and selective manner. Here, we review various aspects of MTase engineering, both methodological and conceptual, and also discuss future directions and challenges. Bacterial MTases that are part of restriction/modification (R/M) systems offer a convenient way for the selection of large gene libraries, both in vivo and in vitro. We review these selection methods, their strengths and weaknesses, and also the prospects for new selection approaches that will enable the directed evolution of mammalian DNA methyltransferases (Dnmts). We explore various properties of MTases that may be subject to engineering. These include engineering for higher stability and soluble expression (MTases, including bacterial ones, are prone to misfolding), engineering of the DNA target specificity, and engineering for the usage of S-adenosyl-L-methionine (AdoMet) analogs. Directed evolution of bacterial MTases also offers insights into how these enzymes readily evolve in nature, thus yielding MTases with a huge spectrum of DNA target specificities. Engineering for alternative cofactors, on the other hand, enables modification of DNA with various groups other than methyl and thus can be employed to map and redirect DNA epigenetic modifications.


Directed Evolution Gene Library Methylation Site Enzyme Variant Consensus Amino Acid 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



DNA methyltransferase


Mammalian DNA methyltransferase




Isopropyl β-D-1-thiogalactopyranoside


In vitro compartmentalization


Polymerase chain reaction


Methylated DNA immunoprecipitation




Nuclear magnetic resonance


Enzyme-linked immunosorbent assay




Sodium dodecyl sulfate-polyacrylamide gel electrophoresis


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Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  • Paola Laurino
    • 1
  • Liat Rockah-Shmuel
    • 1
  • Dan S. Tawfik
    • 1
    Email author
  1. 1.Department of Biological ChemistryWeizmann Institute of ScienceRehovotIsrael

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