Crystal structure and mechanism of action of the N6-methyladenine-dependent type IIM restriction endonuclease R.DpnI.

ABSTRACT: DNA methylation-dependent restriction enzymes have many applications in genetic engineering and in the analysis of the epigenetic state of eukaryotic genomes. Nevertheless, high-resolution structures have not yet been reported, and therefore mechanisms of DNA methylation-dependent cleavage are not understood. Here, we present a biochemical analysis and high-resolution DNA co-crystal structure of the N(6)-methyladenine (m6A)-dependent restriction enzyme R.DpnI. Our data show that R.DpnI consists of an N-terminal catalytic PD-(D/E)XK domain and a C-terminal winged helix (wH) domain. Surprisingly, both domains bind DNA in a sequence- and methylation-sensitive manner. The crystal contains R.DpnI with fully methylated target DNA bound to the wH domain, but distant from the catalytic domain. Ind

SUBMITTER: Siwek W 

PROVIDER: S-EPMC3424567 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature