INTERACTION OF CHOLERAGEN AND THYROTROPIN WITH BOVINE THYROID ADENYLATE-CYCLASE

Activation of adenylate cyclase in a participate fraction of bovine thyroid by choleragen and TSH was studied. Washing the choleragen- or TSH-activated enzyme, followed by incubation at 4 or 37 C for 2–24 h, did not reverse activation. Elution of the TSH-activated enzyme with acidic buffer (pH 2) yielded a substance, presumably TSH, that activated fresh adenylate cyclase. Elution of the choleragen-activated enzyme with acidic buffer yielded a very small amount of material, presumably choleragen, that activated fresh cyclase. Only 3% of choleragen was recoverable, however, from the acid eluate; over 96% of the toxin was inactivated by the thyroid membranes. Incubation of choleragen with the thyroid particulate fraction abolished its ADP-ribosyltransferase activity and considerably reduced its capacity to activate adenylate cyclase. Incubation of TSH with thyroid particulate fraction had no detectable effect, under the conditions used, on its capacity to activate adenylate cyclase. It is concluded that persstent activation of thyroid adenylate cyclase by TSH can be explained by persistent binding of biologically active TSH to the enzyme. There is less evidence that the stability of the choleragen-activated cyclase results from the continued presence of biologically active choleragen. Persistent activation by choleragen may be explained instead by evidence that choleragen activates adenylate cyclase by an NAD+-dependent enzymatic process (Moss, J., and M. Vaughan; JBiol Chem 252: 2455, 1977). The number of cyclase molecules that can be activated by one choleragen molecule may be limited by inactivation of the toxin, which was observed with the thyroid particulate preparation. © 1979 by The Endocrine Society.