Interaction of thyroid peroxidase with concanavalin A covalently coupled to agarose.
We have investigated the interaction between concanavalin A-agarose (Con A-agarose) and thyroid peroxidase, an integral membrane protein found in the 105,000 X g, 1-h particulate fraction of thyroid tissue. An intact form of porcine thyroid peroxidase was obtained by solubilization with the nonionic detergent Triton X-100 and two fragmented, hydrophilic forms of the enzyme were prepared by trypsin treatment of the membrane. The three types of thyroid peroxidase bind to Con A-agarose and can be eluted with alpha-methyl-D-mannoside. The alpha-methyl-D-mannoside eluate of the most purified thyroid peroxidase preparation has been analyzed by polyacrylamide gel electrophoresis. Peroxidase activity corresponds with a glycoprotein band. The binding of thyroid peroxidase to Con A-agarose can be inhibited by sugars in the following order: alpha-methyl-D-mannoside greater than D-mannose greater than alpha-methyl-D-glucoside greater than D-glucose greater than D-galactose. This order of specificity is typical of Con A-sugar interactions. Furthermore, inactivation of the carbohydrate binding site of Con A by demetallization greatly reduces the extent of thyroid peroxidase binding. Reactivation of the carbohydrate binding site by the addition of Ca2+ and Mn2+ to demetallized Con A-agarose restores thyroid peroxidase binding. These and other experiments suggest that htyroid peroxidase is, like several other peroxidases, a glycoprotein. In addition, the interaction between thyroid peroxidase and Con A-agarose may provide a new purification tool for thyroid peroxidase.
D000818:Animals; D001667:Binding, Competitive; D002118:Calcium / Q000494:pharmacology; D002846:Chromatography, Affinity; D003208:Concanavalin A; D004789:Enzyme Activation / Q000187:drug effects; D006601:Hexoses / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007453:Iodide Peroxidase / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D008757:Methylglucosides / Q000494:pharmacology; D008766:Methylmannosides / Q000494:pharmacology; D010544:Peroxidases / Q000378:metabolism; D011485:Protein Binding; D013552:Swine; D013961:Thyroid Gland / Q000201:enzymology
D006601:Hexoses; D008757:Methylglucosides; D008766:Methylmannosides; D003208:Concanavalin A; D008345:Manganese; D010544:Peroxidases; D007453:Iodide Peroxidase; D002118:Calcium