SNAP-annotated 5mC-DNA-protein interactions: PDB entry 4aqx

PDB-id

4aqx; DSSR-derived features in text and JSON formats; DNAproDB

Class

hydrolase

Method

X-ray (2.2 Å)

Summary

Crystal structure of i-crei complexed with its target methylated at position plus 2 (in the b strand) in the presence of magnesium

Reference

Valton J, Daboussi F, Leduc S, Molina R, Redondo P, Macmaster R, Montoya G, Duchateau P (2012): "5'-Cytosine-Phosphoguanine (Cpg) Methylation Impacts the Activity of Natural and Engineered Meganucleases." J.Biol.Chem., 287, 30139. doi: 10.1074/JBC.M112.379966.

Abstract

In this study, we asked whether CpG methylation could influence the DNA binding affinity and activity of meganucleases used for genome engineering applications. A combination of biochemical and structural approaches enabled us to demonstrate that CpG methylation decreases I-CreI DNA binding affinity and inhibits its endonuclease activity in vitro. This inhibition depends on the position of the methylated cytosine within the DNA target and was almost total when it is located inside the central tetrabase. Crystal structures of I-CreI bound to methylated cognate target DNA suggested a molecular basis for such inhibition, although the precise mechanism still has to be specified. Finally, we demonstrated that the efficacy of engineered meganucleases can be diminished by CpG methylation of the targeted endogenous site, and we proposed a rational design of the meganuclease DNA binding domain to alleviate such an effect. We conclude that although activity and sequence specificity of engineered meganucleases are crucial parameters, target DNA epigenetic modifications need to be considered for successful gene editions.

• The contacts include paired nucleotides (mostly a G in Watson-Crick G-C pairing) and amino-acids within a 4.5-Å distance cutoff to base atoms of 5mC.
• The structure is oriented in the base reference frame of 5mC, allowing for easy comparison and direct superimposition between entries.
• The black sphere (•) denotes the 5-methyl carbon atom in 5mC.