title: Purification, crystallization and data collection of the apoptotic nuclease endonuclease G.
abstract: Endonuclease G (EndoG) is a mitochondrial enzyme that responds to apoptotic stimuli by translocating to the nucleus and cleaving chromosomal DNA. EndoG is the main apoptotic endonuclease in the caspase-independent pathway. Mouse EndoG without the mitochondrial localization signal (amino-acid residues 1-43) was successfully overexpressed, purified and crystallized using a microbatch method under oil. The initial crystal (type I) was grown in the presence of the detergent CTAB and diffracted to 2.8 A resolution, with unit-cell parameters a = 72.20, b = 81.88, c = 88.66 A, beta = 97.59 degrees in a monoclinic space group. The crystal contained two monomers in the asymmetric unit, with a predicted solvent content of 46.6%. Subsequent mutation of Cys110 improved the stability of the protein significantly and produced further crystals of types II, III and IV with space groups C2, P4(1)2(1)2 (or P4(3)2(1)2) and P2(1)2(1)2(1), respectively, in various conditions. This suggests the critical involvement of this conserved cysteine residue in the crystallization process.
authors.affiliation: Medical Proteomic Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea.
authors.forename: Sei Mee
authors.initials: SM
authors.lastname: Yoon
authors.forename: Hyung Nam
authors.initials: HN
authors.lastname: Song
authors.forename: Jun Hyuk
authors.initials: JH
authors.lastname: Yang
authors.forename: Mi Yeon
authors.initials: MY
authors.lastname: Lim
authors.forename: Yong Je
authors.initials: YJ
authors.lastname: Chung
authors.forename: Seong Eon
authors.initials: SE
authors.lastname: Ryu
authors.forename: Eui Jeon
authors.initials: EJ
authors.lastname: Woo
mesh_terms: D000818:Animals; D017209:Apoptosis; D003460:Crystallization; D018360:Crystallography, X-Ray; D004706:Endodeoxyribonucleases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice
chemical_list: D004706:Endodeoxyribonucleases; C052391:endonuclease G