Principles of Biochemistry Volume 2 - A. Lehninger 1985

Bioenergetics and Metabolism
Oxidative Degradation of Amino Acids. The Urea Cycle
Chapter Summary

A small fraction of the oxidative energy produced in The Human Body is derived from the oxidative Catabolism of Amino Acids. Following the removal of amino groups in Transamination reactions with α-ketoglutarate, the carbon skeletons of amino acids undergo oxidative degradation and are converted into intermediates that can enter The Citric Acid Cycle and be oxidized to CO₂ and H₂O. There are five pathways by which the carbon skeletons of Amino acids can enter The Citric Acid cycle: 1) via acetyl-CoA, 2) via α-ketoglutarate, 3) via succinate, 4) via fumarate, and 5) via oxaloacetate. Amino acids entering the citric acid cycle via acetyl-CoA are divided into two groups: the first group (Alanine, Cysteine, Glycine, Serine, and Threonine) are converted to acetyl-CoA via Pyruvate, while the second group (leucine, Lysine, phenylalanine, Tyrosine, and Tryptophan) are converted via acetoacetyl-CoA. The carbon skeletons of Proline, Histidine, Arginine, glutamine, and glutamic acid enter the citric acid cycle via α-ketoglutarate; Methionine, isoleucine, and valine enter via succinate; four carbon atoms of Phenylalanine and Tyrosine enter via fumarate; and finally, asparagine and aspartic acid enter via oxaloacetate. In humans, several inborn errors of Amino acid METABOLISM are known. A particularly serious and relatively common disorder of this type is phenylketonuria.

In ammonotelic animals (bony Fishes, tadpoles), amino nitrogen is excreted through the gills as ammonia, which is formed by the Hydrolysis of glutamine. Ureotelic animals (a group that includes most terrestrial animals) excrete amino nitrogen in the form of urea. Urea is synthesized in the Liver; this process,

discovered by Hans Krebs, is known as The Urea Cycle. The immediate precursor of urea is arginine: under the action of arginase, it is hydrolyzed to form urea and Ornithine. Arginine is then resynthesized from ornithine; to achieve this, ornithine is first carbamoylated by carbamoyl phosphate to form citrulline, and then an amino group from aspartate is transferred to citrulline. Ornithine is regenerated in each turn of the urea cycle. Uricotelic animals (birds, snakes, and lizards) excrete amino nitrogen in a semi-solid form as uric acid, which is a purine derivative. The excretion of amino nitrogen as a non-toxic compound (urea) or as a solid (uric acid) requires a significant expenditure of ATP energy by the Organism.

References

Books

Baldwin E. An Introduction to Comparative Biochemistry, 4th ed., Cambridge University Press, New York, 1964.

Cunningham E. B. Biochemistry: Mechanisms of Metabolism, McGraw-Hill, New York, 1978. Chapter 14 provides an excellent description of amino acid metabolic pathways and their enzymatic mechanisms.

Dagley S., Nicholson D. E. An Introduction to Metabolic Pathways, Wiley, New York, 1970. A reference book.

Grisolia S., Baguena R., Mayor F. The Urea Cycle, Wiley, New York, 1976. Proceedings of a symposium on the urea cycle dedicated to the memory of Hans Krebs. Contains interesting material.

Hochachka P. W., Somero G. N. Strategies of Biochemical Adaptation, Holt, Rinehart and Winston, New York, 1973. Comparative biochemistry.

Scriver C. R., Rosenberg L. E. Amino Acid Metabolism and Its Disorders, Saunders, Philadelphia, 1973.

Articles

Holmes F. L., Hans Krebs and the Discovery of the Ornithine Cycle, Fed. Proc., 39, 216-225 (1980). The events leading to the discovery of the cycle, described by a well-known historian of medicine.

Questions and Problems

1. Products of AMINO ACID TRANSAMINATION. Name the α-keto acids formed from the amino acids listed below in the transamination reaction with α-ketoglutarate. Write the structural formulas of these α-keto acids.

a) Aspartic acid.

b) Glutamic acid.

c) Alanine.

d) Phenylalanine.

2. Measurement of the alanine transaminase reaction rate. Alanine transaminase activity (The rate of the alanine transaminase reaction) is typically measured by adding an excess of purified Lactate dehydrogenase and NADH to the reaction system. The rate of alanine disappearance equals the rate of NADH disappearance, which is measured spectrophotometrically. Explain what is happening here.

3. Amino nitrogen distribution. Would you show symptoms of aspartate deficiency on a diet rich in alanine but poor in aspartate? Explain your answer.

4. An inborn error of amino acid metabolism. A two-year-old child is admitted to the hospital. According to the mother, the child suffers from frequent vomiting, occurring mainly after meals. The child shows delayed weight gain and physical development. The Hair is dark, but there are some gray streaks. A urine test after adding FeCl₃ turned green, indicating the presence of phenylpyruvic acid. Quantitative Analysis of the urine yielded the following results:

a) Which enzyme is likely inactive? Suggest a Treatment for this condition.

b) Why does phenylalanine appear in large amounts in the urine?

c) What is the source of phenylpyruvate and phenyllactate? Why does this pathway (which is absent in healthy individuals) begin to function when the phenylalanine concentration rises?

d) Why are there gray streaks in the patient's hair?

5. Role of cobalamin in Amino Acid Catabolism. Pernicious anemia results from an impairment in cobalamin absorption, which is due to the absence of a specific glycoprotein secreted by The Stomach (known as intrinsic factor). How does this impairment affect amino acid catabolism? Does it affect all amino acids equally?

6. Comparison of lactate and alanine as metabolic fuels. ATP energy cost of nitrogen excretion. Based on the oxidation state of the three carbon atoms in lactate and alanine, these compounds are identical; in the animal body, both carbon sources can serve as metabolic fuels. Compare the net ATP yields (number of moles of ATP generated per mole of substrate) for the complete oxidation (to CO₂ and H₂O) of lactate and alanine, taking into account the ATP cost of nitrogen excretion as urea.

Problem 6

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7. Pathway of carbon and nitrogen in glutamate degradation. In rat liver, oxidative degradation occurs for glutamic acid labeled with ¹⁴C at the C-2 position and ¹⁵N in the amino group. A list of metabolites is given below. In which atoms of these metabolites will each label be found?

Problem 7

a) Urea.

b) Succinate.

c) Arginine.

d) Citrulline.

e) Ornithine.

f) Aspartate.

8. Chemical strategy of isoleucine catabolism. Isoleucine is degraded to propionyl-CoA and acetyl-CoA in a six-step pathway.

Problem 8

a) The chemical strategy of isoleucine degradation is analogous to that of the citric acid cycle and the β-Oxidation of Fatty acids. The intermediates of isoleucine degradation (I-V) are shown below out of their metabolic order. Arrange them in the proper metabolic sequence based on your knowledge of the citric acid cycle and Fatty acid oxidation.

b) For each of the steps above, describe the chemical process, identify an analogous reaction in the citric acid cycle or the β-oxidation of Fatty acids, and state the required Cofactors.

9. Ammonia poisoning caused by arginine-free diet. A report [J. Morris, Q. Rogers, Science 199, 431 (1978)] described the following experiment. Cats fasted overnight were given a single meal of an amino acid mixture containing all amino acids except arginine. Within 2 hours, Blood ammonia levels rose to 140 μg/L (normal level is 18 μg/L), and Clinical symptoms of ammonia toxicity appeared. One cat, which consumed only 8 g of the mixture, died within 4.5 hours. In the control group, which received either a complete amino acid mixture or a mixture with arginine replaced by ornithine, no unusual clinical symptoms were observed.

a) What was The Role of the preliminary fasting in this experiment?

b) What is the cause of the elevated blood ammonia level? Why does the absence of arginine in the diet lead to ammonia toxicity? Is arginine an essential amino acid for cats?

c) Why can arginine be replaced by ornithine?

10. Glutamate Oxidation. Write individual balanced equations and the overall equation for glutamate oxidation, in which 2 moles of glutamate yield 2 moles of α-ketoglutarate and 1 mole of urea (excreted).

Electron micrograph of Glycogen granules isolated from rat liver (negative staining). These granules, which represent the storage form of glucose "fuel" in the liver, are called α-particles. They consist of smaller β-particles. The granules contain not only glycogen but also the Enzymes required for its Synthesis and degradation, as well as the enzymes that reciprocally regulate these processes.