Open AccessPurification, crystallization and data collection of the apoptotic nuclease endonuclease G
Author(s) -
Yoon Sei Mee,
Song Hyung Nam,
Yang Jun Hyuk,
Lim Mi Yeon,
Chung Yong Je,
Ryu Seong Eon,
Woo Eui Jeon
Publication year - 2009
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s1744309109013335
Subject(s) - endonuclease , nuclease , crystallization , apoptosis , cysteine , cleavage (geology) , monomer , microbiology and biotechnology , dna , enzyme , chemistry , mutant , monoclinic crystal system , crystallography , biology , biochemistry , crystal structure , gene , paleontology , organic chemistry , fracture (geology) , polymer
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 Å resolution, with unit‐cell parameters a = 72.20, b = 81.88, c = 88.66 Å, β = 97.59° 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 C 2, P 4 1 2 1 2 (or P 4 3 2 1 2) and P 2 1 2 1 2 1 , respectively, in various conditions. This suggests the critical involvement of this conserved cysteine residue in the crystallization process.