title: Crystal structures of type II restriction endonuclease EcoO109I and its complex with cognate DNA.
abstract: EcoO109I is a type II restriction endonuclease that recognizes the DNA sequence of RGGNCCY. Here we describe the crystal structures of EcoO109I and its complex with DNA. A comparison of the two structures shows that the catalytic domain moves drastically to capture the DNA. One metal ion and two water molecules are observed near the active site of the DNA complex. The metal ion is a Lewis acid that stabilizes the pentavalent phosphorus atom in the transition state. One water molecule, activated by Lys-126, attacks the phosphorus atom in an S(N)2 mechanism, whereas the other water interacts with the 3'-leaving oxygen to donate a proton to the oxygen. EcoO109I is similar to EcoRI family enzymes in terms of its DNA cleavage pattern and folding topology of the common motif in the catalytic domain, but it differs in the manner of DNA recognition. Our findings propose a novel classification of the type II restriction endonucleases and lead to the suggestion that EcoO109I represents a new subclass of the EcoRI family.
authors.affiliation: Graduate School of Integrated Science, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hashimoto
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Shimizu
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Imasaki
authors.forename: Matsuri
authors.initials: M
authors.lastname: Kato
authors.forename: Naoki
authors.initials: N
authors.lastname: Shichijo
authors.forename: Keiko
authors.initials: K
authors.lastname: Kita
authors.forename: Mamoru
authors.initials: M
authors.lastname: Sato
mesh_terms: D000595:Amino Acid Sequence; D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004926:Escherichia coli / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D006868:Hydrolysis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D020836:Protein Structure, Quaternary
chemical_list: D004268:DNA-Binding Proteins; D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific; C045450:RGGNCCY-specific type II deoxyribonucleases