Biochemistry 2 Exam 1 (Chapter 14-17)
A researcher adds 14C-labeled glyceraldehyde 3-phosphate to a yeast extract. After a short time, she isolates fructose 1,6-bisphosphate labeled with 14C at C-3 and C-4. What was the location of the 14C label in the starting glyceraldehyde 3-phosphate? Where did the second 14C label in fructose 1,6-bisphosphate come from? Explain.
C-1. This experiment demonstrates the reversibility of the aldolase reaction. The C-1 of glyceraldehyde 3-phosphate is equivalent to C-4 of fructose 1,,6-bisphosphate. The starting glyceraldehyde 3-phosphate must have been labeled at C-1. The C-3 of dihydroxyacetone phosphate becomes labeled through the triose phosphate isomerase reaction, thus giving rise to fructose 1,6-bisphosphate labeled at C-3.
Acyl-CoA dehydrogenase uses enzyme-bound FAD as a prosthetic group to dehydrogenate the alpha and beta carbons of fatty acyl-CoA. What is the advantage of using FAD as an electron acceptor rather than NAD+? Explain in terms of the standard reduction potentials for the Enzyme-FAD/FADH2 (E= -0.219V) and NAD+/NADH (E= -0.320 V) half-reactions.
Enz-FAD, having a more positive standard reduction potential, is a better electron acceptor than NAD+, and the reaction is driven in the direction of fatty acyl-CoA oxidation. This more favorable equilibrium is obtained at the cost of 1 ATPl only 1.5 ATP are produced per FADH2 oxidized in the respiratory chain (vs 2.5 per NADH).
Why is it important that gluconeogenesis is not the exact reversal of glycolysis?
Gluconeogenesis would be highly endergonic, and it would be impossible to separately regulate gluconeogenesis and glycolysis.
Write the net biochemical equation for the metabolism of a molecule of glucose by glycolysis and the citric acid cycle, including all cofactors.
Glucose + 4ADP + 4 Pi + 10NAD+ + 2FAD --> 4ATP + 10NADH + 2FADH2 + 6CO2
For the patient described in the previous question, predict the levels of each listed metabolite in his blood before treatment in the ER, relative to the levels maintained during adequate insulin treatment: (a) glucose (b) ketone bodies (c) free fatty acids
(a) elevated (b) elevated (c) elevated
In muscle tissue, the rate of conversion of glycogen to glucose 6-phosphate is determined by the ratio of phosphorylase a (active) to phosphorylase b (less active). Determine what happens to the rate of glycogen breakdown if a muscle preparation containing glycogen phosphorylase is treated with (a) phosphorylase kinase and ATP (b) PP1 (c) epinephrine.
(a) increases (b) decreases (c) increases
For each of the metabolic transformations, determine whether the compound on the left has undergone oxidation or reduction. (a) Methanol --> formaldehyde (b) Formaldehyde --> formate (c) Carbon dioxide --> formate (d) Glycerate --> glyceraldehyde (e) Glycerol --> dihydroxyacetone (f) Toluene --> benzoate (g) Succinate --> fumarate (h) Pyruvate --> acetate
(a) oxidation (b) oxidation (c) reduction (d) reduction (e) oxidation (f) oxidation (g) oxidation (h) oxidation
Researchers can manipulate genes of a mouse so that a single gene in a single tissue either produces an inactive protein (a "knockout" mouse) or produces a protein that is always active. What effects on metabolism would you predict for mice with the listed genetic changes? (a) Knockout of glycogen debranching enzyme in the liver (b) Knockout of hexokinase IV in the liver (c) Knockout of FBPase-2 in the liver (d) Constitutively active FBPase-2 in liver (e) Constitutively active AMPK in muscle (f) Constitutively active ChREBP in liver
(a) reduced capacity to mobilize glycogen; lowered blood glucose between meals (b) Reduced capacity to lower blood glucose after a carbohydrate meal; elevated blood glucose (c) reduced concentration of fructose 2,6-bisphosphate (F26BP) in liver, stimulating glycolysis and inhibiting gluconeogenesis (d) Reduced [F26BP], stimulating gluconeogenesis and inhibiting glycolysis (e) Increased uptake of fatty acids and glucose; increased oxidation of both (f) Increased conversion of pyruvate to acetyl-CoA; increased fatty acid synthesis
How many turns of the fatty acid oxidation cycle are required for complete oxidation of arachididic acid (20:0) to acetyl-CoA?
9 turns; arachidic acid, a 20-carbon saturated fatty acid, yields 10 molecules of acetyl-CoA, the last two formed in the ninth turn.
When the acetyl‑CoA produced during β oxidation in the liver exceeds the capacity of the citric acid cycle, the excess acetyl‑CoA forms ketone bodies—acetone, acetoacetate, and D‑β‑hydroxybutyrate. This occurs in people with severe, uncontrolled diabetes; because their tissues cannot use glucose, they oxidize large amounts of fatty acids instead. Although acetyl‑CoA is not toxic, the mitochondrion must divert the acetyl‑CoA to ketone bodies. What problem would arise if acetyl‑CoA were not converted to ketone bodies? How does the diversion to ketone bodies solve the problem?
Because the mitochondrial pool of CoA is small, CoA must be recycled from acetyl-CoA via the formation of ketone bodies. This allows the operation of the beta-oxidation pathway, necessary for energy production.
Homeostatic mechanisms maintain the concentration of glucose in human blood at about 5mm. The concentration of free glucose inside a myocyte is much lower. Why is the concentration so low in the cell? What happens to glucose after entry into the cell? Physicians administer glucose intravenously as a food source in certain clinical situations. Given that the transformation of glucose to glucose 6-phosphate consumes ATP, why not administer intravenous glucose 6-phosphate instead?
The phosphate group of glucose 6-phosphate is completely ionized at pH 7, giving the molecule an overall negative charge. Because membranes are generally impermeable to electrically charged molecules, glucose 6-phosphate cannot pass from the bloodstream into cells and hence cannot enter the glycolytic pathway and generate ATP. (This is why glucose, once phosphorylated, cannot escape from the cell.)
Bears expend about 25x10^6 J/day during periods of hibernation, which may last as long as seven months. The energy required to sustain life is obtained from fatty acid oxidation. How much weight (in kg) do bears lose after 7 months of hibernation? How could a bear's body minimize ketosis during hibernation? (Assume the oxidation of fat yields 38 kJ/g)
Mass lost per day is about 0.66 kg or about 140 kg in seven months. Ketosis could be avoided by degradation of nonessential body proteins to supply amino acid skeletons for gluconeogenesis.
The last step of the CAC, NAD+-dependent oxidation of malate forms oxaloacetate. Can a net synthesis of oxaloacetate from Acetyl-CoA occur using only the enzymes and cofactors of the CAC, without depleting the intermediates of the cycle? Explain. How do cells replenish the oxaloacetate that is lost from the cycle to biosynthetic reactions?
No. For every two carbons that enter as acetate, two leave the cycle as CO2; thus there is no net synthesis of oxaloacetate. Net synthesis of oxaloacetate occurs by the carboxylation of pyruvate, an anaplerotic reaction.
During strenuous activity, the demand for ATP in muscle tissue vastly increases. In rabbit leg muscle or turkey flight muscle, ATP production is almost exclusively a product of lactic acid fermentation. Phosphoglycerate kinase and pyruvate kinase catalyze the two reactions that form ATP in the payoff phase of glycolysis. Suppose skeletal muscle were devoid of lactate dehydrogenase. Could it carry out strenuous physical activity; that is, could it generate ATP at a high rate by glycolysis? Explain.
No. Lactate dehydrogenase is required to recycle NAD+ from the NADH formed during the oxidation of glyceraldehyde 3-phosphate.
Suppose you discovered a mutant yeast whose glycolytic pathway was shorter because of the presence of a new enzyme catalyzing the reaction. Glyceraldehyde 3-phosphate + H2O --> 3-phosphoglycerate Would shortening the glycolytic pathway in this way benefit the cell? Explain.
No. There would be no anaerobic production of ATP; aerobic ATP production would be diminished only slightly.
Soy sauce preparation involves fermenting a salted mixture of soybeans and wheat with several microorganisms, including yeast, over a period of 8-12 months. The resulting sauce (after solids are removed) is rich in lactate and ethanol. How are these two compounds produced? To prevent the soy sauce from having a strong vinegary taste (vinegar is dilute acetic acid), oxygen must be kept out of the fermentation tank. Why?
Soybeans and wheat contain starch, a polymer of glucose. The microorganisms break down starch to glucose, glucose to pyruvate via glycolysis, and - because the process is carried out in the absence of O2 - pyruvate to lactate and ethanol. If O2 and H2O were present, some of the acetyl-CoA, however, would also be hydrolyzed to acetic acid (vinegar) in the presence of oxygen.
Two of the steps in the oxidative dexcarboxylation of pyruvate (steps 4 and 5) do not involve any of the three carbons of pyruvate, yet are essential to the operation of the PDH complex. Explain.
Steps 4 and 5 are essential in the reoxidation of the enzymes reduced lipoamide cofactor.
Cells often use the same enzyme reaction pattern for analogous metabolic conversions. For example, the steps in the oxidation of pyruvate to acetyl-CoA and of a-ketoglutarate to succinyl-CoA although catalyzed by different enzymes, are very similar. The first stage of fatty acid oxidation follows a reaction sequence closely resembling a sequence in the Krebs cycle. Use equations to show the analogous reaction sequences in the two pathways.
The first step in fatty acid oxidation is analogous to the conversion of succinate to fumarate; the second step, to the conversion of fumarate to malate; the third step, to the conversion of malate to oxaloacetate.
Citrate is formed by the condensation of acetyl-CoA with oxaloacetate, catalyzed by citrate synthase. Oxaloacetate + acetyl-CoA + H2O --> citrate + CoA + H+ In rat heart mitochondria at pH 7.0 and 25ºC, the concentrations of reactants and products are: oxaloacetate 1 µM; acetyl-CoA 1 µM; citrate 220 µM; and CoA 65 µM. The standard free energy change for the citrate synthase reaction is -32.2 kJ/mol. What is the direction of metabolic flow through the citrate synthase reaction in rat heart cells? Explain.
Toward citrate: delta G for the citrate synthase reaction under these conditions is about -8kJ/mol
What is the structure of the partially oxidized fatty acyl group that is formed when oleic acid (18:1(delta9)), has undergone three cycles of beta oxidation? What are the next two steps in the continued oxidation of this intermediate?
cis-delta3-Dodecanoyl-CoA; it is converted to cis-delta2-dodecanoyl-CoA, then beta-hydroxydodecanoyl-CoA
How many cycles of Beta oxidation are required for the complete oxidation of activated oleic acid, 18:1(delta9)?
8 cycles; the last releases 2 acetyl-CoA
A man with insulin-dependent diabetes is brought to the hospital emergency department in a near-comatose state. While vacationing in an isolated place, he lost his insulin medication and has not taken any insulin in two days. (a) For each tissue listed at the end of the problem, is each pathway faster, slower, or unchanged in this patient, compared with the normal level when he is getting appropriate amounts of insulin? (b) For each pathway, describe at least one control mechanism responsible for the change you predict. Tissue and Pathways: 1. Adipose: fatty acid synthesis 2. Muscle: glycolysis, fatty acid synthesis, glycogen synthesis 3. Liver: glycolysis, gluconeogenesis, glycogen synthesis, fatty acid synthesis, pentose phosphate pathway
(1) Adipose: fatty acid synthesis slower. (2) Muscle: glycolysis, fatty acid synthesis, and glycogen synthesis slower. (3) Liver: glycolysis faster. Gluconeogenesis, glycogen synthesis, and fatty acid synthesis slower. Pentose phosphate pathway unchanged. (b) (1) Adipose and (3) Liver: fatty acid synthesis slower because lack of insulin results in inactive acetyl-CoA carboxylase, the first enzyme of fatty acid synthesis. Glycogen synthesis inhibited by cAMP-dependent phosphorylation (thus activation) of glycogen synthase. (2) Muscle: glycolysis slower because of GLUT4 is inactive, so glucose uptake is inhibited. (3) Liver: glycolysis slower bc of the bifunctional PFK-2/FBPase-2 is converted to the form with active FBPase-2, decreasing [fructose 2,6-bisphosphate], which allosterically stimulates PFK and inhibits FBPase-1; this also accounts for the stimulation of gluconeogenesis.
An investigator carries out a 'pulse-chase' experiment using 14C-labeled carbon sources on yeast extract maintained under strictly anaerobic conditions to produce ethanol. The experiment consists of incubating a small amount of 14C-labeled substrate (the pulse) with the yeast extract just long enough for each intermediate in the fermentation pathway to become labeled. The addition of excess unlabeled glucose then 'chases' the label through the pathway. The chase effectively prevents any further entry of labeled glucose into the pathway. (a) If the investigator uses [1-14C] glucose as a substrate, what is the location of 14C in the product ethanol? (b) Where would 14C have to be located in the starting glucose to ensure that all the 14C activity is liberated as 14CO2 during fermentation to ethanol?
(a) 14CH3CH2OH (b) [3-14C]glucose or [4-14C]glucose
In animal tissues, the ratio of active unphosphorylated to inactive, phosphorylated PDH complex regulates the rate of conversion of pyruvate to acetyl-CoA. Determine what happens to. the rate of this reaction when a preparation of rabbit muscle mitochondria containing the PDH complex is treated with (a) pyruvate dehydrogenase kinase, ATP and NADH (b) pyruvate dehydrogenase phosphatase and Ca2+ (c) malonate
(a) decreases (b) increases (c) decreases
Manufacturers use citric acid as a flavoring agent in soft drinks, fruit juices, and many other foods. Worldwide, the market for citric acid is valued at hundreds of millions of dollars per year. Commercial production uses the mold Aspergillus niger, which metabolizes sucrose under carefully controlled conditions. (a) Why does the yield decrease when the concentration of Fe3 is above or below the optimal value of 0.5 mg/L? (b) Write the sequence of reactions by which A. niger synthesizes citric acid from sucrose. Write an equation for the overall reaction. (c) Does the commercial process require the culture medium to be aerated - that is, is this a fermentation or an anaerobic process? Explain.
(a) Citrate is produced through the action of citrate synthase on oxaloacetate and acetyl-CoA. Citrate synthase can be used for net synthesis of citrate when (1) there is a continuous influx of new oxaloacetate and acetyl-CoA and (2) isocitrate synthesis is restricted, as in a culture medium low in Fe3+. Aconite requires Fe3+, so an Fe3+-restricted medium restricts the synthesis of aconitase. (b) Sucrose + H2O --> glucose + fructose Glucose + 2Pi + 2ADP + 2NAD+ --> 2 pyruvate + 2 ATP + 2NADH + 2H+ + 2H2O Fructose + 2Pi + 2ADP + 2NAD+ --> 2 pyruvate + 2 ATP + 2NADH + 2H+ + 2H2O 2 pyruvate + 2NAD+ + 2CoA --> 2 acetyl-CoA + 2NADH + 2H+ + 2CO2 2 pyruvate + 2CO2 + 2ATP + 2H2O --> 2 oxaloacetate + 2ADP + 2 Pi + 4H+ 2 Acetyl-CoA + 2 oxaloacetate + 2H2O --> 2 citrate + 2CoA Overall: Sucrose + H2O + 2Pi + 2ADP + 6NAD+ --> 2 citrate + 2ATP + 6NADH + 10H+
Suppose you had to subsist on a diet of whale blubber and seal blubber, with little or no carbohydrate. (a) what would be the effect of carbohydrate deprivation on the utilization of fats for energy? (b) if your diet were totally devoid of carbohydrate, would it be better to consume odd- or even-numbered fatty acids? Explain.
(a) Glucose yields pyruvate via glycolysis, and pyruvate is the main source of oxaloacetate. Without glucose in the diet, [oxaloacetate] drops and the citric acid cycle slows. (b) Odd-number, propionate conversion to succinyl-CoA provides intermediates for the citric acid cycle and four-carbon precursors for gluconeogenesis
Predict and explain the effect on glycogen metabolism of each of the listed defects caused by mutation: (a) Loss of the cAMP-binding site on the regulatory subunit of protein kinase A (PKA). (b) Loss of the protein phosphatase inhibitor (c) Over-expression of phosphorylase b kinase in liver (d) Defective glucagon receptors in liver
(a) PKA cannot be activated in response to glucagon or epinephrine, and glycogen phosphorylase is not activated. (b) PP1 remains active, allowing it to dephosphorylate glycogen synthase (activating it) and glycogen phosphorylase (inhibiting it) (c) Phosphorylase remains phosphorylated (active), increasing the breakdown of glycogen (d) Gluconeogenesis cannot be stimulated when blood glucose is low, leading to dangerously low blood glucose during periods of fasting.
An individual developed a condition characterized by progressive muscular weakness and aching muscle cramps. The symptoms were aggravated by fasting, exercise, and a high-fat diet. The homogenate of a skeletal muscle specimen from the patient oxidized added oleate more slowly than did control homogenates, consisting of muscle specimens from healthy individuals. When carnitine was added to the patient's muscle homogenate, the rate of oleate oxidation equaled that in the control homogenates. The patient was diagnosed as having a carnitine deficiency (a) Why did added carnitine increase the rate of oleate oxidation in the patient's muscle homogenate? (b) Why were the patient's symptoms aggravated by fasting, exercise, and a high-fat diet? (c) Suggest two possible reasons for the deficiency of muscle carnitine in this individual.
(a) The carnitine-mediated entry of fatty acids into mitochondria is the rate-limiting step in fatty acid oxidation. Carnitine deficiency slows fatty acid oxidation; added carnitine increases the rate. (b) All increase the metabolic need for fatty acid oxidation. (c) Carnitine deficiency might result from a deficiency of a carnitine precursor (such as lysine), or from a defect in one of the enzymes in the biosynthesis of carnitine.
The graph shows the concentrations of lactate in blood plasma before, during and after a 400m sprint. (a) What causes the rapid rise in lactate concentration? (b) What causes the decline in lactate concentration after completion of the sprint? Why does the decline occur more slowly than the increase? (c) Why is the concentration of lactate not zero during the resting state?
(a) The rapid increase in glycolysis; the rise in pyruvate and NADH results in a rise in lactate. (b) Lactate is transformed to glucose via pyruvate. This is a slower process because formation of pyruvate is limited by NAD+ availability, the lactate dehydrogenase equilibrium is in favor of lactate, and conversion of pyruvate to glucose is energy-requiring. (c) The equilibrium for the lactate dehydrogenase reaction is in favor of lactate formation.
(a) Explain how ATP can be both a substrate and an inhibitor of PFK-1. How is the enzyme regulated by ATP? (b) How do ATP levels regulate glycolysis? (c) The inhibition of PFK-1 by ATP diminishes when the ADP concentration is high, as shown in the graph. What explains this observation?
(a) There are two binding sites for ATP: a catalytic site and a regulatory site. Binding of ATP to a regulatory site inhibits PFK-1, by reducing Vmax or increasing Km for ATP at the catalytic site. (b) Glycolytic flux is reduced when ATP is plentiful. (c) The graph indicates that increased [ADP] suppresses the inhibition by ATP. Because the adenine nucleotide pool is fairly constant, consumption of ATP leads to an increase inn [ADP]. The data show that the activity of PFK-1 may be regulated by the [ATP]/[ADP] ratio.
What is the cost (in ATP equivalents) of transforming glucose to pyruvate via glycolysis and back again to glucose via gluconeogenesis?
4 ATP equivalents per glucose molecule
When grown anaerobically on glucose, yeast (S. cerevisiae) converts pyruvate to acetaldehyde, then reduces acetaldehyde to ethanol using electrons from NADH. Write the equation for the second reaction, and calculate its equilibrium constant at 25*C, given the standard reduction potentials in Table 13-7.
CH3CHO + NADH + H+ <--> CH3CH2OH + NAD+ Keq = 1.45x10^4
Consider the four clinical case studies A through D. For each case determine which enzyme is defective and designate the appropriate treatment from the lists provided at the end of the problem. Justify your choices. Answer the questions contained in each case study. Case A: The patient develops vomiting and diarrhea shortly after milk ingestion. A lactose tolerance test is administered. (The patient ingests a standard amount of lactose, and the blood-plasma glucose levels are measured at intervals. In normal individuals, the levels increase to a maximum in about 1 hour and then recede.) The patient's blood glucose and galactose concentrations do not rise but remain constant. Explain why the blood glucose and galactose increase and then decrease in normal individuals. Why do they fail to rise in the patient? Case B: The patient develops vomiting and diarrhea after ingestion of milk. His blood is found to have a low concentration of glucose but a much higher than normal concentration of reducing sugars. The urine gives a positive test for galactose. Why does galactose appear in the urine? Case C: The patient complains of painful muscle cramps when performing strenuous physical exercise but is otherwise normal. A muscle biopsy indicates that muscle glycogen concentration is much higher than in normal individuals. Why does glycogen accumulate? Case D: The patient is lethargic, her liver is enlarged, and a biopsy of the liver shows large amounts of excess glycogen. She also has a lower than normal level of blood glucose. Account for the low blood glucose concentration in this patient.
Case A: Lactase in intestinal mucosa; Low lactose diet Case B: Galactose 1-phosphate uridylyltransferse; Low lactose diet Case C: Muscle debranching enzyme; avoiding strenuous exercise Case D: Liver glycogen phosphorylase; Frequent feedings (smaller portions) of a normal diet
The consumption of alcohol (ethanol), especially after periods of strenuous activity or after not eating for several hours, results in a deficiency of glucose in the blood, a condition known as hypoglycemia. The first step in the metabolism of ethanol by the liver is oxidation to acetylaldehyde, catalyzed by liver alcohol dehydrogenase. Explain how this reaction inhibits the transformation of lactate to pyruvate. Why does this lead to hypoglycemia?
Consumption of alcohol forces competition for NAD+ between ethanol metabolism and gluconeogenesis. The problem is compounded by strenuous exercise and lack of food, because at these times the level of blood glucose is already low.
The activation of free palmitate to its coenzyme A derivative in the cytosol occurs before it can be oxidized in the mitochondrion. After adding palmitate and [14C] coenzyme A to a liver homogenate, you find pamitatoyl-CoA isolated from the cytosolic fraction is radioactive, but that isolated from the mitochondrial fraction is not. Explain.
Fatty acyl groups condensed with CoA in the cytosol are first transferred to carnitine, releasing CoA, then transported into the mitochondrion, where they are again condensed with CoA. The cytosolic and mitochondrial pools of CoA are thus kept separate, and no radioactive CoA from the cytosolic pool enters the mitochondrion.
In a laboratory experiment, two groups of rats are fed two different fatty acids as their sole source of carbon for a month. The first group gets heptanoic acid (7:0), and the second gets octanoic acid (8:0). After the experiment, those in the first group are healthy and have gained weight, whereas those in the second group are weak and have lost weight as a result of losing muscle mass. What is the biochemical basis for this difference?
For the odd-numbered heptanoic acid, beta oxidation produces proprionyl-CoA, which can be converted in several steps to oxaloacetate, a starting material for gluconeogenesis. The even-numbered fatty acids cannot support gluconeogenesis, bc it is entirely oxidized to acetyl-CoA
A eukaryotic cell can use glucose (C6H12O6) and hexanoate (C6H11O2) as fuels for cellular respiration. On the basis of their structural formulas, which substance releases more energy per gram on complete combustion to CO2 and H2O?
From the structural formulas, we see that the carbon-bound H/C ratio of hexanoate (11/6) is higher than that of glucose (7/6). Hexanoate is more reduced and yields more energy on complete combustion to CO2 and H2O.
The Vmax of the glycogen phosphorylase from skeletal muscle is much greater than the Vmax of the same enzyme from liver tissue. (a) What is the physiological function of glycogen phosphorylase in skeletal muscle? In liver tissue? (b) Why does the Vmax of the muscle enzyme need to be greater than that of the liver enzyme?
In muscle: Glycogen breakdown supplies energy (ATP) via glycolysis. Glycogen phosphorylase catalyzes the conversion of stored glycogen to glucose 1-phosphate, which is converted to glucose 6-phosphate, an intermediate in glycolysis. During strenuous activity, skeletal muscle requires large quantities of glucose 6-phosphate. In the liver: Glycogen breakdown maintains a steady level of blood glucose between meals (glucose 6-phosphate is converted to a free glucose) (b) In actively working muscle, ATP flux requirements are very high and glucose 1-phosphate must be produced rapidly, requiring a high Vmax.
The regulated steps of glycolysis in intact cells can be identified by studying the catabolism of glucose in whole tissues or organs. For example, the glucose consumption by heart muscle can be measured by artificially circulating blood through an isolated intact heart and measuring the concentration of glucose before and after the blood passes through the heart. If the circulating blood is deoxygenated, heart muscle consumes glucose at a steady rate. When oxygen is added to the blood, the rate of glucose consumption drops dramatically, then is maintained at the new, lower rate. Explain.
In the absence of O2, the ATP needs are met by anaerobic glucose metabolism (fermentation to lactate). Because aerobic oxidation of glucose produces far more ATP than does fermentation, less glucose is needed to produce the same amount of ATP.
What type of chemical reaction is involved in the conversion of isocitrate to alpha-ketoglutarate? Name and describe the role of any cofactors. What other reaction(s) of the citric acid cycle are of this same type?
Oxidative decarboxylation; NAD+ or NADP+; alpha-ketoglutarate dehydrogenase reaction
What factors might decrease the pool of oxaloacetate available for the activity of the citric acid cycle? How can the pool of oxaloacetate be replenished?
Oxoloacetate might be withdrawn for aspartate synthesis or for gluconeogenesis. Oxaloacetate is replenished by the anaplerotic reactions catalyzed by PEP carboxykinase, PEP carboxylase, malic enzyme, or pyruvate carboxylase.
Although oxygen does not participate directly in the citric acid cycle, the cycle operates only when O2 is present. Why?
Oxygen is needed to recycle NAD+ from the NADH produced by the oxidative reactions of the citric acid cycle. Reoxidation of NADH occurs during mitochondrial oxidative phosphorylation.
The intracellular use of glucose and glycogen is tightly regulated at four points. To compare the regulation of glycolysis when oxygen is plentiful and when it is depleted, consider the utilization of glucose and glycogen by rabbit leg muscle in two physiological settings: a resting rabbit, with low ATP demands, and a rabbit that sights its mortal enemy, the coyote, and dashes into its burrow. For each setting, determine the relative levels (high, intermediate, or low) of AMP, ATP, citrate, and acetyl-CoA and describe how these levels affect the flow of metabolites through glycolysis by regulating specific enzymes.
Resting: [ATP] high; [AMP] low' [acetyl-CoA] and [citrate] intermediate. Running: [ATP] intermediate; [AMP] high; [acetyl-CoA] and [citrate] low. Glucose flux through glycolysis increases during the anaerobic sprint because (1) the ATP inhibition of glycogen phosphorylase and PFK-1 is partially relieved, (2) AMP stimulates both enzymes, and (3) lower citrate and acetyl-CoA levels relieve their inhibitory effects on PFK-1 and pyruvate kinase, respectively.
Regulation of Citrate Synthase In the presence of saturating amounts of oxaloacetate, the activity of citrate synthase from pig heart tissue shows a sigmoid dependence on the concentration of acetyl-CoA, as shown in the graph below. When succinyl-CoA is added, the curve shifts to the right and the sigmoid dependence is more pronounced. On the basis of these observations, suggest how succinyl-CoA regulates the activity of citrate synthase. Why is succinyl-CoA an appropriate signal for regulation of the citric acid cycle? How does the regulation of citrate synthase control the rate of cellular respiration in pig heart tissue?
Succinyl-CoA is an intermediate of the CAC; its accumulation signals reduced flux through the cycle, calling for reduced entry of acetyl-CoA into the cycle. Citrate synthase, by regulating the primary oxidative pathway of the cell, regulates the supply of NADH and thus the flow of electrons from NADH to O2.
Describe the role of each cofactor involved in the reaction catalyzed by the pyruvate dehydrogenase complex.
TPP: thiazolium ring adds to alpha carbon of pyruvate, then stabilizes the resulting carbanion by acting as an electron sink. Lipoic acid: oxidizes pyruvate to level of acetate (acetyl-CoA) and activates acetate as a thioester. CoA-SH: activates acetate as a thioester. FAD: oxidizes lipoic acid NAD+: oxidizes FADH2
Explain in bioenergetic terms how the conversion of pyruvate to phosphoenolpyruvate in gluconeogenesis overcomes the large, negative, standard free-energy change of the pyruvate kinase reaction in glycolysis.
The cell 'spends' 1 ATP and 1 GTP in converting pyruvate to PEP
The oxidation of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate, catalyzed by glyceraldehyde 3-phosphate dehydrogenase, proceeds with an unfavorable equilibrium constant, yet the flow through this point in the glycolytic pathway proceeds smoothly. How does the cell overcome the unfavorable equilibrium?
The cell rapidly removes the 1,3-bisphosphoglycerate in a favorable subsequent step, catalyzed by phosphoglycerate kinase.
Explain, giving examples, what is meant by the statement that the citric acid cycle is amphibolic.
The cycle participates in catabolic and anabolic processes. For example, it generates ATP by substrate oxidation, but also provides precursors for amino acid synthesis.
Between your evening meal and breakfast, your blood glucose drops and your liver becomes a net producer rather than consumer of glucose. Describe the hormonal basis for this switch, and explain how the hormonal change triggers glucose production by the liver.
The drop in blood glucose triggers release of glucagon by the pancreas. In the liver, glucagon activates glycogen phosphorylase by stimulating its cAMP-dependent phosphorylation and stimulates gluconeogenesis by lowering [fructose 2,6-bisphosphate], thus stimulating FBPase-1.
