Structure and function of the Ah receptor: sulfhydryl groups required for binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin to cytosolic receptor from rodent livers.

Cytosol from rodent liver was exposed to a variety of sulfhydryl-modifying reagents to determine if the cytosolic Ah receptor contained reactive sulfhydryl groups that were essential for preservation of the receptor's ligand binding function. At a 2 mM concentration in rat liver cytosol, all sulfhydryl-modifying reagents tested (except iodoacetamide) both blocked binding of [3H]2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to unoccupied receptor and caused release of [3H]TCDD from receptor sites that had been labeled with [3H]TCDD before exposure to the sulfhydryl-modifying reagent. Exposure of cytosol to iodoacetamide before labeling with [3H]TCDD prevented subsequent specific binding of [3H]TCDD, but iodoacetamide was not effective at displacing previously bound [3H]TCDD from the Ah receptor. The mercurial reagents, mersalyl, mercuric chloride, and p-hydroxymercuribenzoate, were more effective at releasing bound [3H]TCDD from previously labeled sites than were alkylating agents (iodoacetamide, N-ethylmaleimide) or the disulfide compound 5,5'-dithiobis(2-nitrobenzoate). Presence of bound [3H]TCDD substantially protected the Ah receptor against loss of ligand binding function when the cytosol was exposed to sulfhydryl-modifying reagents. This may indicate that the critical sulfhydryl groups lie in or near the ligand binding site on the receptor. Subtle differences exist between the Ah receptor and the receptors for steroid hormones in response to a spectrum of sulfhydryl-modifying reagents, but the Ah receptor clearly contains a sulfhydryl group (or groups) essential for maintaining the receptor in a state in which it can bind ligands specifically and with high affinity.