Chapter 13

Match the terms with the descriptions. (a) Primary messenger (b) Second messenger (c) G-protein coupled receptor (d) Heterotrimeric G-protein (e) \(\mathbf{G}_{\alpha \mathrm{s}}\) (f) Protein kinase \(A\) (g) \(\overline{\text { cAMP }}\) phosphodiesterase (h) GTPase activity (i) Cholera (j) Pertussis (k) Phospholipase \(\mathrm{C}\) (l) \( G_{\alpha q}\) (m) Inositol trisphosphate (n) Protein kinase \(\mathrm{C}\) 1\. Activated by \(7 \mathrm{TM}\) receptor 2\. Stimulated by cAMP 3\. Results in the reassociation of \(G_{\alpha}\) and \(G_{\beta \gamma}\) 4\. Generates two second messengers 5\. Activated by diacylglycerol 6\. Composed of seven transmembrane helices 7\. Message received by the cell 8\. Results from \(G_{\text {ci }}\) inhibition 9\. Activates phospholipase \(\mathrm{C}\) 10 Activates adenylate cyclase 11\. Activates a \(\mathrm{Ca}^{2+}\)channel 12\. Due to persistent stimulation of \(G_{\alpha s}\) 13\. Intracellular chemical that relays message from ligand receptor complex 14\. Results in the inactivation of protein kinase A

What are the three major classes of membrane receptors?

Explain how a small number of hormones binding to the extracellular surface of a cell can have a large biochemical effect inside the cell.

What are some of the structural features common to all membrane-bound receptors?

Why is the GTPase activity of G proteins crucial to the proper functioning of a cell?

Hormones affect the biochemistry of a distinct set of tissues. What accounts for the tissue specificity of hormone action?

Match the terms with the descriptions. (a) Growth hormone binding (b) Growth hormone receptor (c) Receptor tyrosine kinase (d) Grb-2 (e) Sos (f) Ras (g) IRS (h) Phosphoinositide kinase 1 (i) \(\mathrm{PIP}_{3}\) -activated kinase (j) Akt 1\. Dimerization results in cross-phosphorylation 2\. Activates Ras 3\. Activates JAK 4\. Adaptor protein in insulin signaling pathway 5\. Activates Akt 6\. Binds IRS and forms \(\mathrm{IP}_{3}\) 7\. Causes receptor dimerization 8\. Promotes movement of glucose transporters to the cell membrane 9\. Small G-protein 10\. Binds receptor tyrosine kinase and Sos

In an elegant experiment on the nature of receptor tyrosine kinase signaling, a gene was synthesized that encoded a chimeric receptor-the extracellular part came from the insulin receptor, and the membranespanning and cytoplasmic parts came from the EGF receptor. The striking result was that the binding of insulin induced tyrosine kinase activity, as evidenced by rapid autophosphorylation. What does this result tell you about the signaling mechanisms of the EGF and insulin receptors?

An antibody has two identical antigen-binding sites. Remarkably, antibodies to the extracellular parts of growth-factor receptors often lead to the same cellular effects as does exposure to growth factors. Explain this observation.

What is the difference between heterotrimeric G proteins and small G proteins?

A mutated form of the \(\alpha\) subunit of the heterotrimeric G protein has been identified; this form readily exchanges GDP for GTP even in the absence of an activated receptor. What would be the effect on a signaling pathway containing the mutated \(\alpha\) subunit?

Normally, rates of diffusion vary inversely with molecular weights; so smaller molecules diffuse faster than do larger ones. In cells, however, calcium ion diffuses more slowly than does cAMP. Propose a possible explanation.

Glucose is mobilized for ATP generation in muscle in response to epinephrine, which activates \(\mathrm{G}_{\alpha s} .\) Cyclic AMP phosphodiesterase is an enzyme that converts cAMP into AMP. How would inhibitors of cAMP phosphodiesterase affect glucose mobilization in muscle?

Considerable effort has been directed toward determining the genes in which sequence variation contributes to the development of type 2 diabetes, a disease that results from a loss of sensitivity of cells to insulin. Approximately 800 genes have been implicated. Explain the significance of this observation.

Human growth hormone binds to a cell-surface membrane protein that is not a receptor tyrosine kinase. The intracellular domain of the receptor can bind other proteins inside the cell. Furthermore, studies indicate that the receptor is monomeric in the absence of hormone but dimerizes on hormone binding. Propose a possible mechanism for growth-hormone signaling.

Differentiate among a proto-oncogene, an oncogene, and a tumor-repressor gene.

Only one copy of a protooncogene need be mutated to enhance the development of cancer, whereas both copies of a tumor-suppressor gene must be mutated to contribute to cancer development. Explain this distinction.

Because of the high degree of genetic variability in tumors, typically no single anticancer therapy is universally effective for all patients, even within a given tumor type. Hence, it is often desirable to inhibit a particular pathway at more than one point in the signaling cascade. In addition to the EGFR-directed monoclonal antibody cetuximab (p. \(239),\) propose alternative strategies for targeting the EGF signaling pathway for antitumor drug development.

Ligand-gated channels can be thought of as receptors. Explain.

The binding of \(\mathrm{Ca}^{2+}\) to calmodulin induces substantial conformational changes in its EF hands, exposing hydrophobic residues on the surface of the protein. How might this structural change help to propagate the calcium signal?

Suppose that you were investigating a newly discovered growth-factor signal- transduction pathway. You found that, if you added a GTP analog in which the terminal phosphate was replaced by sulfate, the duration of the hormonal response was increased. What can you conclude?

Why is the fact that a monomeric hormone binds to two identical receptor molecules, thus promoting the formation of a dimer of the receptor, considered remarkable?

Some protein kinases are inactive unless they are phosphorylated on key serine or threonine residues. In some cases, active enzymes can be generated by mutating these serine or threonine residues to glutamate. Explain.

At steady state, intracellular levels of \(\mathrm{Ca}^{2+}\) must be kept low to prevent the precipitation of carboxylated and phosphorylated compounds, which form poorly soluble salts with \(\mathrm{Ca}^{2+} .\) The cytoplasmic level of \(\mathrm{Ca}^{2+}\) is approximately \(100 \mathrm{nM},\) several orders of magnitude lower than the concentration in the extracellular medium. How might the cell maintain such low levels of intracellular \(\mathrm{Ca}^{2+}\) ? How does the cell take advantage of the difference in intracellular and extracellular \(\mathrm{Ca}^{2+}\) concentrations?

You prepare a cell line that overexpresses a mutant form of EGFR in which the entire intracellular region of the receptor has been deleted. Predict the effect of overexpression of this construct on EGF signaling in this cell line.

Suppose that each \(\beta\) -adrenergic receptor bound to epinephrine converts 100 molecules of \(G_{\alpha s}\) into their GTP-bound forms and that each molecule of activated adenylate cyclase produces 1000 molecules of \(\mathrm{cAMP}\) per second. With the assumption of a full response, how many molecules of cAMP will be produced in \(1 \mathrm{s}\) after the formation of a single complex between epinephrine and the \(\beta\) -adrenergic receptor?

A scientist wishes to determine the number of receptors specific for a ligand \(\mathrm{X},\) which he has in both radioactive and nonradioactive form. In one experiment, he adds increasing amounts of the radioactive \(\mathrm{X}\) and measures how much of it is bound to the cells. The result is shown as total activity in the adjoining graph. Next, he performs the same experiment, except that he includes a several hundredfold excess of nonradioactive \(\mathrm{X}\). This result is shown as nonspecific binding. The difference between the two curves is the specific binding. (GRAPH CANNOT COPY) (a) Why is the total binding not an accurate representation of the number of receptors on the cell surface? (b) What is the purpose of performing the experiment in the presence of excess nonradioactive ligand? (c) What is the significance of the fact that specific binding attains a plateau?

With the use of experiments such as those described in problems 27 and \(28,\) the number of receptors in the cell membrane can be calculated. Suppose that the specific activity of the radioactive ligand is \(10^{12}\) counts per minute (cpm) per millimole and that the maximal specific binding is \(10^{4}\) cpm per milligram of membrane protein. There are \(10^{10}\) cells per milligram of membrane protein. Assume that one ligand binds per receptor. Calculate the number of receptor molecules present per cell.