4AQU

Crystal structure of I-CreI complexed with its target methylated at position plus 2 (in the b strand) in the presence of calcium

Summary for 4AQU

• Entry DOI: 10.2210/pdb4aqu/pdb
• Related: 1AF5 1BP7 1G9Y 1G9Z 1MOW 1N3E 1N3F 1T9I 1T9J 1U0C 1U0D 2VBJ 2VBL 2VBN 2VBO 4AAB 4AAD 4AAE 4AAF 4AAG
• Descriptor: DNA ENDONUCLEASE I-CREI, 5'-D(*DTP*CP*AP*AP*AP*AP*CP*GP*TP*CP*GP*TP*GP*DAP *GP*AP*CP*AP*GP*TP*TP*TP*GP*G)-3', 5'-D(*DCP*CP*AP*AP*AP*CP*TP*GP*TP*CP*TP*CP*AP*5CMP *GP*AP*CP*GP*TP*TP*TP*TP*GP*A)-3', ... (6 entities in total)
• Functional Keywords: hydrolase, gene targeting, protein-dna interaction, homing endonucleases
• Biological source: CHLAMYDOMONAS REINHARDTII; More
• Cellular location: Plastid, chloroplast: P05725
• Total number of polymer chains: 4
• Total formula weight: 50049.16

Authors

Valton, J.,Daboussi, F.,Leduc, S.,Redondo, P.,Macmaster, R.,Molina, R.,Montoya, G.,Duchateau, P. (deposition date: 2012-04-19, release date: 2012-07-04, Last modification date: 2023-12-20)

Primary citation

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

PubMed 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.

PubMed: 22740697
DOI: 10.1074/JBC.M112.379966

Experimental method

X-RAY DIFFRACTION (2.3 Å)