Crystallization and molecular-replacement studies of the monoclonal antibody mAbR310 specific for the (R)-HNE-modified protein.
4-hydroxy-2-nonenal (HNE), a major racemic product of lipid peroxidation, reacts with histidine to form a stable HNE-histidine Michael addition-type adduct possessing three chiral centres in the cyclic hemiacetal structure. Monoclonal antibodies against HNE-modified protein have been widely used for assessing oxidative stress in vitro and in vivo. Here, the purification, crystallization and preliminary crystallographic analysis of a Fab fragment of novel monoclonal antibody R310 (mAbR310), which recognizes (R)-HNE-modified protein, are reported. The Fab fragment of mAbR310 was obtained by digestion with papain, purified and crystallized. Using hanging-drop vapour-diffusion crystallization techniques, crystals of mAbR310 Fab were obtained. The crystal belongs to the monoclinic space group C2 (unit-cell parameters a = 127.04, b = 65.31, c = 64.29 A, beta = 118.88 degrees ) and diffracted X-rays to a resolution of 1.84 A. The asymmetric unit contains one molecule of mAbR310, with a corresponding crystal volume per protein weight of 2.51 A(3) Da(-1) and a solvent content of 51.0%.
Ito
Sohei
S
Department of Food and Nutritional Sciences, Graduate School of Nutritional and Environmental Sciences, University of Shizuoka, 52-1 Yada, Suruga-Ku, Shizuoka 422-8526, Japan. itosohei@u-shizuoka-ken.ac.jp
Tatsuda
Emi
E
Ishino
Kousuke
K
Suzuki
Kenichiro
K
Sakai
Hiroshi
H
Uchida
Koji
K
D000447:Aldehydes / Q000737:chemistry*; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D003460:Crystallization; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry / Q000276:immunology*; D012997:Solvents; D014961:X-Ray Diffraction
D000447:Aldehydes; D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D011506:Proteins; D012997:Solvents; C462359:tert-4-hydroxy-2-nonenal