Zheng, YuCohen-Karni, DevoraXu, DerrickChin, Hang GyeongWilson, GeoffreyPradhan, SriharsaRoberts, Richard J.2012-01-122012-01-1220102010-5-5Wang, Yong, Eric A. Franzosa, Xiang-Sun Zhang, Yu Xia. "A unique family of Mrr-like modification-dependent restriction endonucleases" Nucleic Acids Research 38(16): 5527-5534. (2010)1362-4962https://hdl.handle.net/2144/3367Mrr superfamily of homologous genes in microbial genomes restricts modified DNA in vivo. However, their biochemical properties in vitro have remained obscure. Here, we report the experimental characterization of MspJI, a remote homolog of Escherichia coli's Mrr and show it is a DNA modification-dependent restriction endonuclease. Our results suggest MspJI recognizes mCNNR (R = G/A) sites and cleaves DNA at fixed distances (N12/N16) away from the modified cytosine at the 3′ side (or N9/N13 from R). Besides 5-methylcytosine, MspJI also recognizes 5-hydroxymethylcytosine but is blocked by 5-glucosylhydroxymethylcytosine. Several other close homologs of MspJI show similar modification-dependent endonuclease activity and display substrate preferences different from MspJI. A unique feature of these modification-dependent enzymes is that they are able to extract small DNA fragments containing modified sites on genomic DNA, for example ^∼32 bp around symmetrically methylated CG sites and ^∼31 bp around methylated CNG sites. The digested fragments can be directly selected for high-throughput sequencing to map the location of the modification on the genomic DNA. The MspJI enzyme family, with their different recognition specificities and cleavage properties, provides a basis on which many future methods can build to decode the epigenomes of different organisms.enCopyright Wang, Yong, Eric A. Franzosa, Xiang-Sun Zhang, Yu Xia 2010. Published by Oxford University Press.http://creativecommons.org/licenses/by-nc/2.5Article10.1093/nar/gkq327204448792938202