title: Visualizing phosphodiester-bond hydrolysis by an endonuclease.
abstract: The enzymatic hydrolysis of DNA phosphodiester bonds has been widely studied, but the chemical reaction has not yet been observed. Here we follow the generation of a DNA double-strand break (DSB) by the Desulfurococcus mobilis homing endonuclease I-DmoI, trapping sequential stages of a two-metal-ion cleavage mechanism. We captured intermediates of the different catalytic steps, and this allowed us to watch the reaction by 'freezing' multiple states. We observed the successive entry of two metals involved in the reaction and the arrival of a third cation in a central position of the active site. This third metal ion has a crucial role, triggering the consecutive hydrolysis of the targeted phosphodiester bonds in the DNA strands and leaving its position once the DSB is generated. The multiple structures show the orchestrated conformational changes in the protein residues, nucleotides and metals during catalysis.
authors.affiliation: Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Molina
authors.affiliation: 1] Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain. [2] Macromolecular Crystallography Group, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Stefano
authors.initials: S
authors.lastname: Stella
authors.affiliation: Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.
authors.forename: Pilar
authors.initials: P
authors.lastname: Redondo
authors.affiliation: Joint Barcelona Computing Center (BSC)-Centre for Genomic Regulation (CRG)-Institute for Research in Biomedicine (IRB) Program in Computational Biology, Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain.
authors.forename: Hansel
authors.initials: H
authors.lastname: Gomez
authors.affiliation: Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.
authors.forename: María José
authors.initials: MJ
authors.lastname: Marcaida
authors.affiliation: 1] Joint Barcelona Computing Center (BSC)-Centre for Genomic Regulation (CRG)-Institute for Research in Biomedicine (IRB) Program in Computational Biology, Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain. [2] Departament de Bioquimica, Facultat de Biologia, University of Barcelona, Barcelona, Spain.
authors.forename: Modesto
authors.initials: M
authors.lastname: Orozco
authors.affiliation: Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.
authors.forename: Jesús
authors.initials: J
authors.lastname: Prieto
authors.affiliation: 1] Macromolecular Crystallography Group, Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain. [2] Macromolecular Crystallography Group, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Guillermo
authors.initials: G
authors.lastname: Montoya
mesh_terms: D004247:DNA / Q000378:metabolism*; D019620:Desulfurococcaceae / Q000201:enzymology*; D004720:Endonucleases / Q000378:metabolism*; D004952:Esters / Q000378:metabolism*; D006868:Hydrolysis; D008958:Models, Molecular; D010710:Phosphates / Q000378:metabolism*; D011487:Protein Conformation
chemical_list: D004952:Esters; D010710:Phosphates; D004247:DNA; D004720:Endonucleases