High affinity thyroid hormone-binding protein in human kidney: kinetic characterization and identification by photoaffinity labeling.

The binding of thyroid hormones and its regulation of NADPH and NADP+ were studied in human kidney cytosol, and a 38-kDa polypeptide (p38) was identified by photoaffinity labeling of cytosol with underivatized [125I]T3, SDS-PAGE, and autoradiography. The cytosolic thyroid hormone binding and p38 photolabeling were strongly activated by NADPH (maximum at 10(-7) M), whereas other nucleotides were less effective or ineffective. NADP+ did not activate T3 binding and p38 photolabeling, provided it was protected from conversion to NADPH by the addition of an exogenous oxidizing enzymatic system (oxidized glutathione plus glutathione reductase). Furthermore, NADP+ inhibited NADPH activation (half-maximum inhibitory effect at approximately 2 x 10(-5)M), and oxidation of NADPH to NADP+ induced dissociation of bound T3. The equilibrium dissociation constant (Kd) of the NADPH-activated cytosolic T3-binding sites was 0.3 nM, similar to the Kd of the nuclear T3 receptors. The kidney contained 200 times more cytosolic ...