Transient complexes. A structural model for the activation of adenylate cyclase by hormone receptors (guanine nucleotides/irradiation inactivation)

Abstract

1. The irradiation-inactivation procedure was used to study changes in the state of association of the protein components of adenylate cyclase in intact rat liver plasma membranes by measurement of alterations in the target size determined from the catalytic activity of the enzyme. 2. A decrease in target size at 30 degrees C in response to p[NH]ppG (guanosine 5'-[betagamma-imido]triphosphate) or GTP was demonstrated, which we take to reflect the dissociation of a regulatory subunit. The effect of GTP is potentiated by glucagon. This effect is not observed at 0 degrees C. 3. An increase in target size was observed in response to glucagon in the absence of guanine nucleotides, which we take to reflect the association of glucagon receptor with adenylate cyclase. 4. We propose a model for the activation of adenylate cyclase by glucagon in which the binding of the hormone to its receptor causes an initial association of the receptor with the catalytic unit of the enzyme and a regulatory subunit to form a ternary complex. The subsequent activation of the adenylate cyclase results from the dissociation of the ternary complex to leave a free catalytic unit in the activated state. This dissociation requires the binding of a guanine nucleotide to the regulatory subunit. 5. The effects of variation of temperature on the activation of adenylate cyclase by glucagon and guanine nucleotides were examined and are discussed in relation to the irradiation-activation data. 6. The effectiveness of hormones, guanine nucleotides and combinations of hormone and guanine nucleotides as activators of adenylate cyclase in both rat liver and rat fat-cell plasma membranes was studied and the results are discussed in relation to the model proposed, which is also considered in relation to the observations published by other workers.

Mechanism of glucagon activation of adenylate cyclase in the presence of Mn2+

“…Hence, under these conditions, hormone-stimulated activity is relatively insensitive to receptor loss [14]. In contrast, the addition of glucagon alone appears to promote a mobile receptor mechanism of coupling [13,14,23]. Here, the receptor is presumed to form a ~ransmembrane complex with an activated catalytic unit and regulatory protein (see [14]).…”

“…Only in the presence of hormone do they interact physically to elicit activation of adenylate cyclase. The basic difference in these mechanisms is that under conditions where activation via a mobile receptor model occurs, then these 3 interacting entities form a stable, activated, transmembrane complex, However, when activation occurs via a collision coupling mechanism, this complex breaks down to release a transiently activated catalytic unit [13][14][15]231.…”

“…[ 12,15,16] and neurohypophyseal hormone [ 171 is achieved via productive collisions, within the plane of the membrane, between these various components. In liver, 3 independent methods (receptor depletion [14]; Arrhenius plot analysis [14]; irradiation inactivation [ 13]), have been used to support this contention.…”

“…The guanine nucleotide regulatory proteins appear to regulate the coupling interaction between the receptor and catalytic unit [9-121. There is now considerable evidence consistent with the hypothesis that hormonal activation of adenylate cyclase by glucagon [ 11,13,14], &agonists Here, we examine the effect of replacing Mg2+ with Mn2+, in the adenylate cyclase assay, on the activity of the enzyme in liver plasma membranes and its mode of coupling to the glucagon receptor.…”

Insulin controls the cyclic AMP‐dependent phosphorylation of integral and peripheral proteins associated with the rat liver plasma membrane

“…That obtained for cyclic AMP is rather intriguing, as basal intracellular cyclic AMP levels in the liver are likely to be 0.3-0.5 PM [ 19,201: too low to allow these phosphorylation events to occur. However, after stimulation of liver cells by glucagon, cyclic AMP levels rise to 2-4 I.IM [21,22]. This would allow the phosphorylation of the integral proteins to ensue and would allow the phosphorylation of the peripheral proteins to occur if insulin was present at an appropriate concentration.…”