G Protein–Coupled Receptors and Second Messengers:-The-Adrenergic Receptor System Acts through the Second Messenger cAMP

Epinephrine action begins when the hormone binds to a protein receptor in the plasma membrane of a hormone sensitive cell. Adrenergic receptors (“adrenergic” reflects the alternative name for epinephrine, adrenaline) are of four general types, α1, α2, β1, and β2, defined by subtle differences in their affinities and responses to a group of agonists and antagonists. Agonists are structural analogs that bind to a receptor and mimic the effects of its natural ligand; antagonists are analogs that bind without triggering the normal effect and thereby block the effects of agonists. In some cases, the affinity of the synthetic agonist or antagonist for the receptor is greater than that of the natural agonist (Fig. 12–11). The four types of adrenergic receptors are found in different target tissues and mediate different responses to epinephrine. Here we focus on the β -adrenergic receptors of muscle, liver, and adipose tissue. These receptors mediate changes in fuel metabolism, including the increased break down of glycogen and fat. Adrenergic receptors of the β 1 and β 2 subtypes act through the same mechanism, so in our discussion, “β -adrenergic” applies to both types.

The β -adrenergic receptor is an integral protein with seven hydrophobic regions of 20 to 28 amino acid residues that “snake” back and forth across the plasma membrane seven times. This protein is a member of a very large family of receptors, all with seven transmembrane helices, that are commonly called serpentine receptors, G protein–coupled receptors (GPCR), or 7 transmembrane segment (7tm) receptors. The binding of epinephrine to a site on the receptor deep within the membrane (Fig. 12–12, step 1) promotes a conformational change in the receptor’s intracellular domain that affects its interaction with the second protein in the signal-transduction pathway, a heterotrimeric GTP-binding stimulatory G protein, or G_s, on the cytosolic side of the plasma membrane. Alfred G. Gilman and Martin Rodbell discovered that when GTP is bound to G_s, G_s stimulates the production of cAMP by adenylyl cyclase (see below) in the plasma membrane. The function of Gs as a molecular switch resembles that of another class of G proteins typified by Ras, discussed in Section 12.3 in the context of the insulin receptor. Structurally, Gs and Ras are quite distinct; G proteins of the Ras type are monomers (Mr ∼20,000), whereas the G proteins that interact with serpentine receptors are trimers of three different subunits, α (Mr 43,000), β (Mr 37,000), and γ (Mr 7,500 to 10,000).

FIGURE 12–11 Epinephrine and its synthetic analogs. Epinephrine, also called adrenaline, is released from the adrenal gland and regulates energy-yielding metabolism in muscle, liver, and adipose tissue. It also serves as a neurotransmitter in adrenergic neurons. Its affinity for its receptor is expressed as a dissociation constant for the receptor-ligand complex. Isoproterenol and propranolol are synthetic analogs, one an agonist with an affinity for the receptor that is higher than that of epinephrine, and the other an antagonist with extremely high affinity.

When the nucleotide-binding site of Gs (on the subunit) is occupied by GTP, Gs is active and can activate adenylyl cyclase (AC in Fig. 12–12); with GDP bound to the site, Gs is inactive. Binding of epinephrine enables the receptor to catalyze displacement of bound GDP by GTP, converting Gs to its active form (step 2). As this occurs, the β and γ subunits of Gs dissociate from the α subunit, and Gsα, with its bound GTP, moves in the plane of the membrane from the receptor to a nearby molecule of adenylyl cyclase (step 3 ). The Gs is held to the membrane by a covalently attached palmitoyl group (see Fig. 11–14). Adenylyl cyclase (Fig. 12–13) is an integral protein of the plasma membrane, with its active site on the cytosolic face. It catalyzes the synthesis of cAMP from ATP:

The association of active G_sα with adenylyl cyclase stimulates the cyclase to catalyze cAMP synthesis (Fig. 12–12, step 4), raising the cytosolic [cAMP]. This stimulation by G_sα is self-limiting; G_sα is a GTPase that turns itself off by converting its bound GTP to GDP (Fig. 12–14). The now inactive G_sα dissociates from adenylyl cyclase, rendering the cyclase inactive. After Gs reassociates with the β and γ subunits (G_{s β γ}), Gs is again available to interact with a hormone-bound receptor.

FIGURE 12–12 Transduction of the epinephrine signal: the β-adrenergic pathway. The seven steps of the mechanism that couples binding of epinephrine (E) to its receptor (Rec) with activation of adenylyl cyclase (AC) are discussed further in the text. The same adenylyl cyclase molecule in the plasma membrane may be regulated by a stimulatory G protein (Gs), as shown, or an inhibitory G protein (Gi, not shown). Gs and Gi are under the influence of different hormones. Hormones that induce GTP binding to Gi cause inhibition of adenylyl cyclase, resulting in lower cellular [cAMP].

FIGURE 12–13 Interaction of G_sα with adenylyl cyclase. (PDB ID 1AZS) The soluble catalytic core of the adenylyl cyclase (AC, blue), severed from its membrane anchor, was cocrystallized with G_sα (green) to give this crystal structure. The plant terpene forskolin (yellow) is a drug that strongly stimulates the enzyme, and GTP (red) bound to G_sα triggers interaction of G_sα with adenylyl cyclase.

FIGURE 12–14 Self-inactivation of Gs. The steps are further described in the text. The protein’s intrinsic GTPase activity, in many cases stim ulated by RGS proteins (regulators of G protein signaling), determines how quickly bound GTP is hydrolyzed to GDP and thus how long the G protein remains active.

FIGURE 12–15 Activation of cAMP-dependent protein kinase, PKA. (a) A schematic representation of the inactive R2C2 tetramer, in which the autoinhibitory domain of a regulatory (R) subunit occupies the substrate-binding site, inhibiting the activity of the catalytic (C) sub unit. Cyclic AMP activates PKA by causing dissociation of the C sub units from the inhibitory R subunits. Activated PKA can phosphorylate a variety of protein substrates (Table 12–3) that contain the PKA consensus sequence (X–Arg–(Arg/Lys)–X–(Ser/Thr)–B, where X is any residue and B is any hydrophobic residue), including phosphorylase b kinase. (b) The substrate-binding region of a catalytic subunit revealed by x-ray crystallography (derived from PDB ID 1JBP). Enzyme side chains known to be critical in substrate binding and specificity are in blue. The peptide substrate (red) lies in a groove in the enzyme surface, with its Ser residue (yellow) positioned in the catalytic site. In the inactive R₂C₂ tetramer, the autoinhibitory domain of R lies in this groove, blocking access to the substrate.

One downstream effect of epinephrine is to activate glycogen phosphorylase b. This conversion is promoted by the enzyme phosphorylase b kinase, which catalyzes the phosphorylation of two specific Ser residues in phosphorylase b, converting it to phosphorylase a (see Fig. 6–31). Cyclic AMP does not affect phosphorylase b ki nase directly. Rather, cAMP-dependent protein ki nase, also called protein kinase A or PKA, which is allosterically activated by cAMP (Fig. 12–12, step 5), catalyzes the phosphorylation of inactive phosphorylase b kinase to yield the active form. The inactive form of PKA contains two catalytic sub units (C) and two regulatory subunits (R) (Fig. 12–15a), which are similar in sequence to the catalytic and regulatory domains of PKG (cGMP-dependent protein ki nase). The tetrameric R2C2 complex is catalytically in active, because an autoinhibitory domain of each R subunit occupies the substrate-binding site of each C subunit. When cAMP binds to two sites on each R sub unit, the R subunits undergo a conformational change and the R₂C₂ complex dissociates to yield two free, catalytically active C subunits. This same basic mechanism—displacement of an autoinhibitory domain— mediates the allosteric activation of many types of protein kinases by their second messengers (as in Figs 12–7 and 12–23, for example).

As indicated in Figure 12–12 (step 6 ), PKA regu lates a number of enzymes (Table 12–3). Although the proteins regulated by cAMP-dependent phosphorylation have diverse functions, they share a region of sequence similarity around the Ser or Thr residue that undergoes phosphorylation, a sequence that marks them for regulation by PKA. The catalytic site of PKA (Fig. 12–15b) interacts with several residues near the Thr or Ser residue in the target protein, and these interactions de fine the substrate specificity. Comparison of the sequences of a number of protein substrates for PKA has yielded the consensus sequence—the specific neigh boring residues needed to mark a Ser or Thr residue for phosphorylation (see Table 12–3). Signal transduction by adenylyl cyclase entails several steps that amplify the original hormone signal (Fig. 12–16). First, the binding of one hormone molecule to one receptor catalytically activates several Gs molecules. Next, by activating a molecule of adenylyl cyclase, each active Gs molecule stimulates the catalytic synthesis of many molecules of cAMP. The second messenger cAMP now activates PKA, each molecule of which catalyzes the phosphorylation of many molecules of the target protein—phosphorylase b kinase in Figure 12–16. This kinase activates glycogen phosphorylase b, which leads to the rapid mobilization of glucose from glycogen. The net effect of the cascade is amplification of the hormonal signal by several orders of magnitude, which accounts for the very low concentration of epinephrine (or any other hormone) required for hormone activity. Cyclic AMP, the intracellular second messenger in this system, is short-lived. It is quickly degraded by cyclic nucleotide phosphodiesterase to 5-AMP (Fig. 12–12, step 7), which is not active as a second messenger:

The intracellular signal therefore persists only as long as the hormone receptor remains occupied by epinephrine. Methyl xanthines such as caffeine and theophylline (a component of tea) inhibit the phosphodiesterase, increasing the half-life of cAMP and thereby potentiating agents that act by stimulating adenylyl cyclase.

FIGURE 12–16 Epinephrine cascade. Epinephrine triggers a series of reactions in hepatocytes in which catalysts activate catalysts, resulting in great amplification of the signal. Binding of a small number of molecules of epinephrine to specific β-adrenergic receptors on the cell surface activates adenylyl cyclase. To illustrate amplification, we show 20 molecules of cAMP produced by each molecule of adenylyl cyclase, the 20 cAMP molecules activating 10 molecules of PKA, each PKA molecule activating 10 molecules of the next enzyme (a total of 100), and so forth. These amplifications are probably gross underestimates.

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مواضيع ذات صلة

Solute Transport across Membranes:-The β-Adrenergic Receptor Is Desensitized by Phosphorylation

Receptor Enzymes:- Receptor Guanylyl Cyclases Generate the Second Messenger cGMP

Receptor Enzymes: -The Insulin Receptor Is a Tyrosine-Specific Protein Kinase

Gated Ion Channels:- Neurons Have Receptor Channels That Respond to Different Neurotransmitters

Gated Ion Channels:- Voltage-Gated Ion Channels Produce Neuronal Action Potentials

Gated Ion Channels:- Ion Channels Underlie Electrical Signaling in Excitable Cells

Solute Transport across Membranes:- Defective Ion Channels Can Have Adverse Physiological Consequences

Solute Transport across Membranes:- The Acetylcholine Receptor Is a Ligand-Gated Ion Channel

Solute Transport across Membranes:- The Neuronal Na+ Channel Is a Voltage-Gated Ion Channel

Solute Transport across Membranes: -The Structure of a K+ Channel Reveals the Basis for Its Specificity

Solute Transport across Membranes:- Ion-Channel Function Is Measured Electrically

Solute Transport across Membranes:- Ion-Selective Channels Allow Rapid Movement of Ions across Membranes