Biochemical characterization of selenium-containing catalytic antibody as a cytosolic glutathione peroxidase mimic.
A selenium-containing catalytic antibody (Se-4A4), prepared by converting reactive serine residues of a monoclonal antibody (4A4) raised against a GSH derivative into selenocysteines, acts as a mimic of cytosolic glutathione peroxidase (cGPX). To clarify the mechanism of action of this catalytic antibody, detailed studies on kinetic behaviour and biological activity were carried out. A rate of acceleration (kcat/Km/kuncat) 10(7)-fold that of the uncatalytic reaction is observed. Under similar conditions, the turnover number (kcat) of Se-4A4 is 42% of that of the natural rabbit liver cGPX. The Se-4A4 reaction involves a Ping Pong mechanism, which is the same as that of the natural cGPX. The selenocysteine residue is located in the binding site of the antibody and is shown to be crucial for this activity. Of the thiol compounds tested, only GSH is able to serve as substrate for Se-4A4. It was demonstrated, using the free-radical-damage system (hypoxanthine/xanthine oxidase) of cardiac mitochondria, that Se-4A4 can protect mitochondria from free-radical damage at least 10(4)-fold more effectively than the natural cGPX.
Ding
L
L
Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, People's Republic of China.
Liu
Z
Z
Zhu
Z
Z
Luo
G
G
Zhao
D
D
Ni
J
J
D017151:Antibodies, Catalytic / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D005609:Free Radicals / Q000009:adverse effects / Q000378:metabolism; D005978:Glutathione / Q000276:immunology / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D019271:Hypoxanthine / Q000378:metabolism; D007700:Kinetics; D015227:Lipid Peroxidation / Q000502:physiology; D008929:Mitochondria, Heart / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology; D011506:Proteins / Q000378:metabolism*; D012643:Selenium / Q000032:analysis; D051140:Selenoproteins; D013379:Substrate Specificity; D014969:Xanthine Oxidase / Q000378:metabolism
D017151:Antibodies, Catalytic; D000911:Antibodies, Monoclonal; D005609:Free Radicals; D011506:Proteins; D051140:Selenoproteins; D019271:Hypoxanthine; D006861:Hydrogen Peroxide; D005979:Glutathione Peroxidase; D014969:Xanthine Oxidase; D005978:Glutathione; D012643:Selenium