title: Kinetics of oxidation of o-dianisidine by hydrogen peroxide in the presence of antibody complexes of iron(III) coproporphyrin.
abstract: The complex of iron(III) coproporphyrinI (FeCPI) with antibody D5E3 was studied as an artificial peroxidase, using o-dianisidine as a substrate. At saturation with respect to antibody, the initial rates of o-dianisidine oxidation are practically the same for free and bound FeCPI at a concentration 5 x 10(-9)M, but the catalytic rate constant (kc) for bound FeCPI exceed (kc) for free FeCPI by two- to three-fold. This difference can be explained by a real enhancement of (kc) at the antibody-active site. The dependence of initial rates of the reaction on substrate concentrations obeyed Michaelis-Menten kinetics and revealed substrate activation at high concentrations of o-dianisidine. A comparison of the Stern-Volmer constants for o-dianisidine-induced quenching of the porphyrin fluorescence proves that antibody-bound coproporphyrin is equivalently accessible to the substrate as protoporphyrin bound to apoperoxidase from horseradish peroxidase (HRP). Based on analysis of the (kc) dependence on H2O2 concentrations in the FeCPI-antibody system, we suggest that interaction with hydrogen peroxide is the rate-limiting step for the oxidation reaction.
authors.affiliation: Laboratory of Molecular Immunology, A. N. Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow.
authors.forename: A P
authors.initials: AP
authors.lastname: Savitsky
authors.forename: M I
authors.initials: MI
authors.lastname: Nelen
authors.forename: A K
authors.initials: AK
authors.lastname: Yatsmirsky
authors.forename: M V
authors.initials: MV
authors.lastname: Demcheva
authors.forename: G V
authors.initials: GV
authors.lastname: Ponomarev
authors.forename: I V
authors.initials: IV
authors.lastname: Sinikov
mesh_terms: D017151:Antibodies, Catalytic / Q000276:immunology / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D002384:Catalysis; D003306:Coproporphyrins / Q000378:metabolism*; D003962:Dianisidine / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism*; D007700:Kinetics; D010084:Oxidation-Reduction; D013050:Spectrometry, Fluorescence
chemical_list: D017151:Antibodies, Catalytic; D000911:Antibodies, Monoclonal; D003306:Coproporphyrins; C089596:iron coproporphyrin; D006861:Hydrogen Peroxide; D003962:Dianisidine