An engineered split M.HhaI-zinc finger fusion lacks the intended methyltransferase specificity.
The ability to site-specifically methylate DNA in vivo would have wide applicability to the study of basic biomedical problems as well as enable studies on the potential of site-specific DNA methylation as a therapeutic strategy for the treatment of diseases. Natural DNA methyltransferases lack the specificity required for these applications. Nomura and Barbas [W. Nomura, C.F. Barbas 3rd, In vivo site-specific DNA methylation with a designed sequence-enabled DNA methylase, J. Am. Chem. Soc. 129 (2007) 8676-8677] have reported that an engineered DNA methyltransferase comprised of fragments of M.HhaI methyltransferase and zinc finger proteins has very high specificity for the chosen target site. Our analysis of this engineered enzyme shows that the fusion protein methylates target and non-target sites with similar efficiency.
Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, MD 21218, USA.
Glenna E
GE
Meister
Srinivasan
S
Chandrasegaran
Marc
M
Ostermeier
D004247:DNA / Q000737:chemistry; D019175:DNA Methylation; D004262:DNA Restriction Enzymes / Q000737:chemistry; D015257:DNA-Cytosine Methylases / Q000737:chemistry* / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics; D017422:Sequence Analysis, DNA; D013447:Sulfites / Q000737:chemistry; D016335:Zinc Fingers
D011993:Recombinant Fusion Proteins; D013447:Sulfites; D004247:DNA; C037664:DNA modification methylase HhaI; D015257:DNA-Cytosine Methylases; D004262:DNA Restriction Enzymes; C042345:hydrogen sulfite