Purification of TSH receptor from porcine thyroid membrane and effect of various protease inhibitors on receptor stability.
Purification of porcine thyroid TSH receptors was attempted by using Lubrol, TSH-Sepharose, gel filtration and the immunoabsorption by anti-thyroglobulin (Tg) antibody-Sepharose. The effects of microbial protease inhibitors on the degradation of TSH receptors were also evaluated. Porcine thyroid membrane was solubilized with 1% Lubrol-PX and TSH receptors were purified by TSH affinity chromatography, followed by gel filtration on TSKgel-G3000SW. A partial purification was achieved; a 692-fold purification and 49.3% recovery of high affinity binding site and a 409-fold purification and 29.1% recovery of low affinity binding site, respectively. Rechromatography on G3000SW gel and immunoabsorption by anti-Tg antibody-Sepharose resulted in further reduction of contaminants. Among 10 protease inhibitors studied, a low concentration of chymostatin was the only one that showed a significant protective effect on the degradation of the receptors (p less than 0.05).
Akamizu
T
T
Department of Medicine, Kyoto University Faculty of Medicine, Japan.
Mori
T
T
Ishii
H
H
Yotota
T
T
Nakamura
H
H
Imura
H
H
D000818:Animals; D001323:Autoantibodies; D002462:Cell Membrane / Q000032:analysis / Q000648:ultrastructure; D002846:Chromatography, Affinity; D007163:Immunosorbent Techniques; D011480:Protease Inhibitors / Q000494:pharmacology*; D011989:Receptors, Thyrotropin / Q000302:isolation & purification* / Q000378:metabolism; D013552:Swine; D013961:Thyroid Gland / Q000648:ultrastructure*; D013972:Thyrotropin / Q000378:metabolism
D001323:Autoantibodies; D011480:Protease Inhibitors; D011989:Receptors, Thyrotropin; C006606:anti-thyroglobulin; D013972:Thyrotropin