Chapter 27

(Integrates with Chapters 3,18 , and \(22 .\) ) The conversion of PEP to pyruvate by pyruvate kinase (glycolysis) and the reverse reaction to form PEP from pyruvate by pyruvate carboxylase and PEP carboxykinase (gluconeogenesis) represent a so-called substrate cycle. The direction of net conversion is determined by the relative concentrations of allosteric regulators that exert kinetic control over pyruvate kinase, pyruvate carboxylase, and PEP carboxykinase. Recall that the last step in glycolysis is catalyzed by pyruvate kinase: \(P E P+A D P \rightleftharpoons\) pyruvate \(+\) ATP The standard free energy change is \(-31.7 \mathrm{kJ} / \mathrm{mol}\). a. Calculate the equilibrium constant for this reaction. b. If \([\mathrm{ATP}]=[\mathrm{ADP}],\) by what factor must [pyruvate] exceed [PEP] for this reaction to proceed in the reverse direction? The reversal of this reaction in eukaryotic cells is essential to gluconeogenesis and proceeds in two steps, each requiring an equivalent of nucleoside triphosphate energy: c. The \(\Delta G^{\circ}\) ' for the overall reaction is \(+0.8 \mathrm{kJ} /\) mol. What is the value of \(K_{\mathrm{eq}} ?\) d. Assuming [ATP] = [ADP], [GTP] = [GDP], and Pi \(=1 \mathrm{m} M\) when this reaction reaches equilibrium, what is the ratio of \([\mathrm{PEP}] /[\text { pyruvate }]\) e. Are both directions in the substrate cycle likely to be strongly favored under physiological conditions?

Strenuous muscle exertion (as in the 100 -meter dash) rapidly depletes ATP levels. How long will 8 m \(M\) ATP last if 1 gram of muscle consumes \(300 \mu\) mol of ATP per minute? (Assume muscle is \(70 \%\) water.) Muscle contains phosphocreatine as a reserve of phosphorylation potential. Assuming [phosphocreatine] \(=40 \mathrm{m} M,[\text { creatine }]=4 \mathrm{m} M,\) and \(\left.\Delta G^{\circ \prime} \text { (phosphocreatine }+\mathrm{H}_{2} \mathrm{O} \rightleftharpoons \text { creatine }+\mathrm{P}_{\mathrm{i}}\right)=-43.3 \mathrm{kJ} / \mathrm{mol}\) how low must [ATP] become before it can be replenished by the reaction: phosphocreatine \(+\mathrm{ADP} \rightleftharpoons \mathrm{ATP}+\) creatine? [Remember \(\Delta G^{\circ \prime}\) (ATP hydrolysis) \(=-30.5 \mathrm{kJ} / \mathrm{mol} .\)]

(Integrates with Chapters \(18 \text { and } 22 .)\) The reactions catalyzed by PFK and FBPase constitute another substrate cycle. PFK is AMP activated; FBPase is AMP inhibited. In muscle, the maximal activity of PFK (mmol of substrate transformed per minute) is ten times greater than FBPase activity. If the increase in [AMP] described in problem 5 raised PFK activity from \(10 \%\) to \(90 \%\) of its maximal value but lowered FBPase activity from \(90 \%\) to \(10 \%\) of its maximal value, by what factor is the flux of fructose- 6 - \(P\) through the glycolytic pathway changed? (Hint: Let PFK maximal activity = 10, FBPase maximal activity \(=1 ;\) calculate the relative activities of the two enzymes at low \([\mathrm{AMP}]\) and at high \([\mathrm{AMP}] ;\) let \(J,\) the flux of \(\mathrm{F}\) - 6 -P through the substrate cycle under any condition, equal the velocity of the PFK reaction minus the velocity of the FBPase reaction.)

(Integrates with Chapters 23 and 24 .) Leptin not only induces synthesis of fatty acid oxidation enzymes and uncoupling protein 2 in adipocytes, but it also causes inhibition of acetyl-CoA carboxylase, esulting in a decline in fatty acid biosynthesis. This effect on acetyl CoA carboxylase, as an additional consequence, enhances fatty acid oxidation. Explain how leptin- induced inhibition of acetyl-CoA carboxylase might promote fatty acid oxidation.

(Integrates with Chapters 19 and \(20 .\) ) Acetate produced in ethanol metabolism can be transformed into acetyl-CoA by the acetyl thiokinase reaction: $$\text { Acetate }+\mathrm{ATP}+\mathrm{CoASH} \longrightarrow \text { acetyl-CoA }+\mathrm{AMP}+\mathrm{PP}_{\mathrm{i}}$$ Acetyl-CoA then can enter the citric acid cycle and undergo oxidation to \(2 \mathrm{CO}_{2}\). How many ATP equivalents can be generated in a liver cell from the oxidation of one molecule of ethanol to \(2 \mathrm{CO}_{2}\) by this route, assuming oxidative phosphorylation is part of the process? (Assume all reactions prior to acetyl-CoA entering the citric acid cycle occur outside the mitochondrion.) Per carbon atom, which is a better metabolic fuel, ethanol or glucose? That is, how many ATP equivalents per carbon atom are generated by combustion of glucose versus ethanol to \(\mathrm{CO}_{2}\) ?

(Integrates with Chapter \(23 .\) ) Assuming each NADH is worth 3 ATP, each \(\mathrm{FADH}_{2}\) is worth \(2 \mathrm{ATP}\), and each NADPH is worth \(4 \mathrm{ATP}\) How many ATP equivalents are produced when one molecule of palmitoyl-CoA is oxidized to 8 molecules of acetyl-CoA by the fatty acid \(\beta\) -oxidation pathway? How many ATP equivalents are consumed when 8 molecules of acetyl-CoA are transformed into one molecule of palmitoyl-CoA by the fatty acid biosynthetic pathway? Can both of these metabolic sequences be metabolically favorable at the same time if \(\Delta G\) for ATP synthesis is \(+50 \mathrm{kJ} / \mathrm{mol}\) ?

The existence of leptin was revealed when the ob/ob genetically obese strain of mice was discovered. These mice have a defective leptin gene. Predict the effects of daily leptin injections into \(o b / o b\) mice on food intake, fatty acid oxidation, and body weight. Similar clinical trials have been conducted on humans, with limited success. Suggest a reason why this therapy might not be a miracle cure for overweight individuals.

Would it be appropriate to call neuropeptide \(Y\) (NPY) the obesitypromoting hormone? What would be the phenotype of a mouse whose melanocortin-producing neurons failed to produce melanocortin? What would be the phenotype of a mouse lacking a functional MC3R gene? What would be the phenotype of a mouse lacking a functional leptin receptor gene?

The Human Biochemistry box, The Metabolic Effects of Alcohol Consumption, points out that ethanol is metabolized to acetate in the liver by alcohol dehydrogenase and aldehyde dehydrogenase: $$\begin{array}{r} \mathrm{CH}_{3} \mathrm{CH}_{2} \mathrm{OH}+\mathrm{NAD}^{+} \rightleftharpoons \mathrm{CH}_{3} \mathrm{CHO}+\mathrm{NADH}+\mathrm{H}^{+} \\\ \mathrm{CH}_{3} \mathrm{CHO}+\mathrm{NAD}^{+}+\mathrm{H}_{2} \mathrm{O} \rightleftharpoons \mathrm{CH}_{3} \mathrm{COO}^{-}+\mathrm{NADH}+2 \mathrm{H}^{+} \end{array}$$ These reactions alter the NAD \(^{+} /\) NADH ratio in liver cells. From your knowledge of glycolysis, gluconeogenesis, and fatty acid oxidation, what might be the effect of an altered \(\mathrm{NAD}^{+} / \mathrm{NADH}\) ratio on these pathways? What is the basis of this effect?

a. Leptin was discovered when a congenitally obese strain of mice \((o b / o b \text { mice })\) was found to lack both copies of a gene encoding a peptide hormone produced mainly by adipose tissue. The peptide hormone was named leptin. Leptin is an anorexic (appetitesuppressing agent; its absence leads to obesity. Propose an experiment to test these ideas. b. A second strain of obese mice \((d b / d b\) mice ) produces leptin in abundance but fails to respond to it. Assuming the \(d b\) mutation leads to loss of function in a protein, what protein is likely to be nonfunctional or absent? How might you test your idea?