Interaction of lactoperoxidase with thiols and diiodotyrosine.
Glutathione and cysteine bind to the heme of lactoperoxidase, thereby causing a red shift of the Soret band which is reversed upon addition of iodide or guaiacol, two substrates for lactoperoxidase. The rate of formation of the enzyme-thiol complex is enhanced by diiodotyrosine. Binding of diiodotyrosine to lactoperoxidase does not cause a shift of the Soret band which indicates binding to the protein of the enzyme. At neutral pH and low ionic strength, lactoperoxidase is adsorbed on insolubilized diiodotyrosine (diiodotyrosine-agarose). It can be eluted at slightly increased ionic strength which shows that the binding is weak. In the presence of 5 X 10(-4) M glutathione, however, the binding of the enzyme to diiodotyrosine-agarose becomes much stronger so that a high salt concentration is required for elution. Lactoperoxidase is also adsorbed on insolubilized thiols (thiol-agarose). The presence of diiodotyrosine is not required for strong binding. A simple method for the preparation of lactoperoxidase from milk by affinity chromatography is based on the interactions of the enzyme with the two ligands, thiols and diiodotyrosine.
D003545:Cysteine; D004105:Diiodotyrosine; D005978:Glutathione; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007784:Lactoperoxidase / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism; D011485:Protein Binding; D013053:Spectrophotometry
D004105:Diiodotyrosine; D007784:Lactoperoxidase; D010544:Peroxidases; D005978:Glutathione; D003545:Cysteine