Human Biochemistry, Volume 1 - Murray R. 1993

Metabolism of Proteins and Amino Acids
Catabolism of the Carbon Skeletons of Amino Acids
Amino Acids Forming Succinyl-CoA

Overall Reactions

During the catabolic breakdown of Methionine, valine, and isoleucine into the amphibolic end product succinyl-CoA, a portion of the carbon Skeleton is lost (Fig. 31.21). Four of the five carbon atoms of valine, three of the five carbon atoms of methionine, and half of the carbon atoms of isoleucine are converted into a succinyl-CoA molecule. The carboxyl carbon atoms of all these Amino Acids yield CO₂. The two terminal carbon atoms of the isoleucine molecule form acetyl-CoA, and the S-methyl group of methionine is removed.

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Fig. 31.21. Summary Scheme for the Catabolism of methionine, isoleucine, and valine converted to succinyl-CoA. As-CoA — acetyl-CoA.

The following describes only the reactions that lead to The conversion of methionine and isoleucine into propionyl-CoA and the conversion of valine into methylmalonyl-CoA. The reactions converting propionyl-CoA to methylmalonyl-CoA and further to succinyl-CoA were already discussed in Chapter 23 in connection with the catabolism of propionate and odd-chain Fatty acids.

Methionine

Interaction of L-methionine with ATP yields S-adenosylmethionine ("active methionine") (Fig. 31.22). The activated S-methyl group can subsequently be transferred to a variety of acceptor compounds¹. Removal of the methyl group yields S-adenosylhomocysteine. Hydrolysis of the S—C bond produces L-homocysteine and adenine. Homocysteine then condenses with Serine to form cystathionine (Fig. 31.23). Hydrolytic Cleavage of cystathionine yields L-homoserine and Cysteine, so the overall process results in the conversion of homocysteine to homoserine and serine to cysteine. These same two reactions are involved in The Biosynthesis of cysteine from serine (see Chapter 29). Homoserine is converted to a-ketobutyrate through the action of homoserine deaminase (Fig. 31.24). This is followed by the conversion of a-ketobutyrate to propionyl-CoA, which proceeds via the conventional pathway of oxidative decaroxylation of a-keto acids (Pyruvate, a-ketoglutarate) to form acyl-CoA derivatives.

Metabolic Disorders of methionine catabolism. See Table 31.2.

¹ Compounds that receive a methyl group from S-adenosylmethionine include betaines, Choline, creatine, epinephrine, melatonin, Sarcosine, N-methylated amino acids, NUCLEOTIDES, and many plant Alkaloids.

Fig. 31.22. Formation of S-adenosylmethionine. ~CH₃ represents the "active methionine" group with a high transfer potential.

Fig. 31.23. Conversion of methionine to propionyl-CoA.

Fig. 31.24. Conversion of L-homoserine to a-ketobutyrate catalyzed by homoserine deaminase.

Leucine, Valine, and Isoleucine

As might be expected given the structural similarities of L-leucine, L-valine, and L-isoleucine, their catabolism shares a common initial pathway. Subsequently, this pathway diverges, and the skeleton of each amino acid is transformed via its own specific route to yield amphibolic intermediates (Figs. 31.25 and 31.26). Depending on The Nature of these amphibolic end products, Amino acids are classified as glucogenic (valine), ketogenic (leucine), or both (isoleucine). Many of the reactions under consideration are analogous to the catabolism of straight-chain and Branched-Chain Fatty Acids. Because the initial catabolic steps for all Three amino acids are similar—as illustrated in Fig. 31.26—it is convenient to discuss them together. In the text that follows, reaction numbers correspond to those given in Figs. 31.26–31.29.

A. Transamination. The reversible transamination (Reaction 1) of all three branched-chain L-a-amino acids in mammalian Tissues is presumably carried out by a single transaminase. The reversibility of this reaction explains why dietary L-a-Amino acids can be replaced by their corresponding a-keto acids, provided adequate sources of nitrogen are available to the Organism.

B. Oxidative decarboxylation yielding acyl-CoA thioesters. This reaction (Reaction 2) is analogous to the Oxidation of Pyruvate to acetyl-CoA and CO₂ by pyruvate dehydrogenase and The oxidation of a-ketoglutarate to CO₂ and succinyl-CoA by a-ketoglutarate dehydrogenase (see Chapter 18). In mammals, the branched-chain a-keto acid dehydrogenase is a mitochondrial multienzyme complex that catalyzes The oxidative decarboxylation of a-ketoisocaproate (from leucine), a-keto-ß-methylvalerate (from isoleucine), and a-ketoisovalerate (from valine).

The subunits of the a-keto acid dehydrogenase complex are analogous to the corresponding subunits of pyruvate dehydrogenase. The complex includes subunits with a-keto acid decarboxylase, transacylase, and dihydrolipoyl dehydrogenase activities. As with pyruvate dehydrogenase, this complex is inactivated by ATP-dependent phosphorylation in a reaction catalyzed by a protein kinase. Ca²⁺-independent phosphoprotein phosphatase catalyzes the dephosphorylation of the complex and thereby reactivates it. Thus, Conversion of the enzyme to the phosphorylated state can regulate branched-chain Amino Acid Catabolism. The protein kinase is inhibited by ADP, branched-chain a-keto acids, the hypolipidemic agent clofibrate, dichloroacetate, and coenzyme A thioesters (e.g., acetoacetyl-CoA). Among the branched-chain a-keto acids, a-ketoisocaproate (a-ketoleucine) is the most potent inhibitor.

Fig. 31.25. Catabolism of Branched-chain amino acids in mammals. Reactions 1–3 are common to all three amino acids, after which their Catabolic pathways diverge. Crossed-out arrows indicate steps where METABOLISM is blocked in two rare Genetic Disorders: 2 — maple syrup urine disease (catabolism of all three amino acids is impaired), 3 — isovaleric acidemia (leucine catabolism is impaired).

C. Dehydrogenation yielding a,ß-unsaturated acyl-CoA thioesters. This reaction (Reaction 3) is analogous to the dehydrogenation of straight-chain acyl-CoA thioesters during fatty acid catabolism. It remains unclear whether the dehydrogenation of all three branched-chain acyl-CoA thioesters is catalyzed by the same dehydrogenase. Indirect evidence points to the involvement of at least two Enzymes, based on examinations of patients with isovaleric acidemia who accumulate isovalerate in their Blood following a protein-rich meal, while levels of other branched-chain a-keto acids remain normal. Isovalerate is formed by the deacylation of isovaleryl-CoA, which is the substrate for the aforementioned dehydrogenase.

Reactions Specific to Leucine Catabolism (Fig. 31.27)

Reaction 4L: Carboxylation of ß-methylcrotonyl-CoA. A key observation that explained the ketogenic effect of leucine was the discovery that the conversion of leucine's terminal isopropyl group to acetoacetate involves the "fixation" (i.e., covalent attachment) of 1 mole of CO₂ per mole of isopropyl groups. This fixation (Reaction 4L, Fig. 31.27) is preceded by the ATP-driven addition of CO₂ to enzyme-bound biotin to form biotinyl-CO₂. As a result of CO₂ fixation, the intermediate ß-methylglutaconyl-CoA is formed.

Fig. 31.26. Similarities in the first Three Reactions of leucine, valine, and isoleucine catabolism. Note that reactions 2 and 3 are also analogous to the corresponding fatty acid catabolism reactions. A similar analogy is evident at several subsequent stages (see subsequent figures).

Reaction 5L: addition of Water across the double bond of ß-methylglutaconyl-CoA. The product of this reaction, ß-hydroxy-ß-methylglutaryl-CoA, serves as a precursor not only for Ketone Bodies (reaction 6L, Fig. 31.27) but also for mevalonate, which can be further converted into Cholesterol and other polyisoprenoids (see Ch. 28).

Reaction 6L: cleavage of ß-hydroxy-ß-methylglutaryl-CoA. The reaction in which acetyl-CoA and acetoacetate are formed from ß-hydroxy-ß-methylglutaryl-CoA takes place in the Cell/35.html">Mitochondria of Liver, Kidney, and Heart Cells in mammals. This process accounts for the strong ketogenic effect of leucine, since 1 mole of leucine yields, In addition to 1 mole of acetoacetate, an indirect formation of another 1/2 mole of ketone bodies from acetyl-CoA (see Ch. 28).

Reactions specific to valine catabolism (Fig. 31.28)

Reaction 4V: Hydration of methacrylyl-CoA. This reaction, which proceeds fairly rapidly even in the absence of an enzyme, is catalyzed by crotonase, a broadly specific hydrolase acting on L-ß-hydroxyacyl-CoA thioesters containing 4 to 9 carbon atoms in the oxyacyl moiety.

Reaction 5V: deacylation of ß-hydroxyisobutyryl-CoA. Since the CoA thioester is not a substrate for the subsequent reaction (reaction 6V, Fig. 31.28), it must first be deacylated to ß-hydroxyisobutyrate (reaction 5V, Fig. 31.28). This reaction is catalyzed by a deacylase present in many animal tissues; the deacylase can also act on another substrate, ß-hydroxypropionyl-CoA.

Reaction 6V: oxidation of ß-hydroxyisobutyrate. Mammalian tissues contain an enzyme that catalyzes the NAD⁺-dependent oxidation of the primary alcohol group of ß-hydroxyisobutyrate to an aldehyde group (reaction 6V, Fig. 31.28), yielding methylmalonic semialdehyde. The reaction is readily reversible.

Fig. 31.27. Catabolism of ß-methylcrotonyl-CoA derived from L-leucine (see Fig. 31.26). Asterisks denote carbon atoms derived from CO₂. The Structure of biotinyl-CO₂ is shown in Fig. 20.4.

Reaction 7V: further transformations of methylmalonate semialdehyde. Subsequent metabolism of methylmalonate semialdehyde in mammalian tissues can proceed via two pathways: transamination yields ß-aminoisobutyrate (reaction 7V, Fig. 31.28), whereas the pathway depicted in Fig. 31.28 (reactions 8V–10V) leads to succinyl-CoA. Transamination to form a-aminoisobutyrate—an amino acid normally present in urine—takes place in various mammalian tissues, including the Kidneys. The second major pathway involves oxidation to methylmalonate, followed by The formation of methylmalonyl-CoA and its subsequent isomerization into succinyl-CoA (reactions 8V–10V, Fig. 31.28). Isomerization (reaction 10V, Fig. 31.28) proceeds with the participation of the adenosylcobalamin coenzyme and is catalyzed by methylmalonyl-CoA mutase. This reaction is crucial for the catabolism not only of valine but also of the propionyl-CoA catabolite derived from isoleucine (Fig. 31.29). Under conditions of cobalamin (vitamin B₁₂) deficiency, mutase activity decreases; in ruminants that utilize propionate produced by rumen Fermentation as an energy source, this manifests as a "nutritional metabolic defect." Purified mutase from sheep liver contains 2 molecules of deoxyadenosyl-vitamin B₁₂ per enzyme molecule. Conversion to succinyl-CoA occurs via intramolecular transfer of the CoA-linked carboxyl group. Although the overall reaction resembles the isomerization of threo-ß-methylaspartate to glutamate, the mechanisms of these reactions likely differ.

Reactions specific to isoleucine catabolism (Fig. 31.29)

Much like in the case of valine and leucine, the earliest insights into isoleucine catabolism were gained through observations of animals fed various diets, which revealed both the glycogenic and weakly ketogenic properties of isoleucine. The possibility of Glycogen synthesis from isoleucine was confirmed using D₂O. Experiments with ¹⁴C-labeled compounds demonstrated that the carbon skeleton of isoleucine is cleaved to yield acetyl-CoA and propionyl-CoA (Fig. 31.29).

Reaction 4I: hydration of tiglyl-CoA. This reaction proceeds analogously to the valine catabolism reaction (reaction 4V, Fig. 31.28) and is catalyzed by mammalian tissue crotonase.

Fig. 31.28. Catabolism of methacrylyl-CoA derived from L-valine (see Fig. 31.26). a-KA — a-keto acid, a-AA — a-amino acid.

Reaction 5I: dehydrogenation of a-methyl-ß-hydroxybutyryl-CoA. This reaction is analogous to reaction 5V of valine catabolism (Fig. 31.28). Recall that during valine catabolism, the corresponding acyl-CoA is first hydrolyzed and only subsequently oxidized.

Reaction 6I: thiolysis of a-methylacetoacetyl-CoA. The thiolytic Cleavage of the covalent bond between carbon atoms 2 and 3 in the a-methylacetoacetyl-CoA molecule is catalyzed by ß-ketothiolase, similarly to the thiolysis of acetoacetyl-CoA to form two molecules of acetyl-CoA. The Nature of the resulting products—acetyl-CoA (ketogenic) and propionyl-CoA (glycogenic)—accounts for the ketogenic and glycogenic properties of isoleucine.

Inborn errors of branched-chain amino acid catabolism (leucine, valine, and isoleucine)

Four metabolic disorders are currently known. The most thoroughly studied is maple syrup urine disease (named after the characteristic odor of the patients' urine), with over 50 cases described. Its incidence is estimated at 5 to 10 per million newborns. The other three disorders—hypervalinemia, intermittent ketonuria (elevated levels of branched-chain keto acids in the urine), and isovaleric acidemia—have each been observed in fewer than five children.

Fig. 31.29. Catabolism of tiglyl-CoA derived from L-isoleucine (see Fig. 31.26).

A. Hypervalinemia. This metabolic disorder is characterized by elevated plasma levels of valine (but not leucine or isoleucine), resulting from a defect in the transamination of valine to form a-ketoisovalerate (reaction 1, Fig. 31.26). Meanwhile, the transamination of leucine and isoleucine (reaction 1, Fig. 31.26) remains unimpaired.

B. Maple syrup urine disease. The most striking hallmark of this inherited disorder is the distinctive odor of the patient's urine, resembling maple syrup or burnt sugar. Plasma and urine show markedly elevated concentrations of the branched-chain amino acids—leucine, isoleucine, and valine—as well as their corresponding a-keto acids. For this reason, maple syrup urine disease is sometimes referred to as branched-chain ketoaciduria. The presence of branched-chain a-hydroxy acids, formed via the reduction of a-keto acids, has also been detected in the urine.

The characteristic symptoms of the disease manifest by the end of the first week of life. Alongside the biochemical abnormalities described above, feeding difficulties arise, and vomiting may occur. Lethargy is sometimes observed. Diagnosing the condition prior to the end of the first week is possible only through Enzymatic Analysis. Affected children who survive exhibit profound neurological impairment. In the absence of Treatment, the disease proves fatal by the end of the first year of life.

The biochemical cause of the disease is the absence or a severe decrease in The activity of branched-chain $$α-keto acid decarboxylase, which catalyzes the conversion of all three branched-chain $$α-keto acids into acyl-CoA thioesters with the release of CO₂ (reaction 2, Fig. 31.26). This was established through enzymatic analysis of leukocytes and cultured Skin fibroblast cells from affected children. The Mechanism of the Toxic Effect of the accumulating compounds remains unknown.

Patients are prescribed a diet replacing Proteins with a mixture of purified amino acids free of leucine, isoleucine, and valine. Once the levels of these three amino acids in the plasma drop to normal, they can be reintroduced into the diet—for example, as part of milk and other foods—but only in amounts sufficient to meet (without exceeding) the requirements for branched-chain amino acids. There is no clear data on whether or when dietary restrictions can subsequently be eased. If treatment is initiated within the first week of life, the severe manifestations of the disease can be significantly mitigated.

B. Intermittent branched-chain ketonuria. This disorder is a variant of maplesyrup urine disease and is presumably associated with relatively minor structural alterations in the α-keto acid decarboxylase. The decarboxylase activity in leukocytes and fibroblasts is significantly lower than normal, yet substantially higher than in classic cases of maple-syrup urine disease. Because patients in this category retain a reduced but still measurable capacity for the catabolism of leucine, isoleucine, and valine, the Typical symptoms of maple-syrup urine disease manifest later and occur only episodically. The prognosis is more favorable with appropriate dietary management.

Maple-syrup urine disease and intermittent branched-chain ketonuria serve to illustrate the principle discussed in the Introduction to this chapter: Mutations that cause varying structural modifications in the same enzyme. Presumably, differences in enzyme activity are reflected across a spectrum of disease manifestations, ranging from a severe phenotype and intermittent ketonuria to a virtually normal state.

C. Isovaleric acidemia. The clinical manifestations of this disorder include a cheesy odor of the breath and Body Fluids, vomiting, acidosis, and even coma, the latter of which may be precipitated by excessive protein intake or secondary infections. In three documented cases, moderate mental retardation was observed. The underlying cause of the disease is a deficiency of isovaleryl-CoA dehydrogenase (reaction 3, Fig. 31.26). The accumulating isovaleryl-CoA undergoes hydrolysis, and the resulting isovalerate is excreted in the urine and sweat.

Additional metabolic disorders associated with amino acid catabolism (propionate, methylmalonate, and vitamin B₁₂)

Propionyl-CoA (Fig. 31.21) can be derived from isoleucine (Figs. 31.26 and 31.29), methionine (Fig. 31.23), as well as from the side chain of cholesterol and odd-chain fatty acids. The conversion of propionyl-CoA into amphibolic intermediates involves a biotin-dependent carboxylation to yield methylmalonyl-CoA; the latter can also be formed directly from valine (without The intermediate formation of propionyl-CoA) (Figs. 31.21 and 31.28, reaction 9 V). Through an isomerization reaction co-enzymed by a vitamin B₁₂ derivative, methylmalonyl-CoA is converted into succinyl-CoA—an intermediate of The Citric Acid Cycle that is further oxidized to CO₂ and water.

Shortly after the involvement of 5'-deoxyadenosylcobalamin as a cofactor in the isomerization of methylmalonyl-CoA to succinyl-CoA was established, it was demonstrated that patients with vitamin B₁₂ deficiency excrete large amounts of methylmalonate in the urine. Methylmalonic aciduria ceased following the administration of adequate amounts of vitamin B₁₂.

A. Propionic acidemia. Deficient propionyl-CoA carboxylase activity is characterized by elevated serum propionate levels and impaired propionate catabolism in leukocytes. Treatment consists of a restricted diet and measures to mitigate metabolic acidosis.

B. Methylmalonic aciduria. Two forms of methylmalonic aciduria are known. One of these responds to parenteral administration of physiological doses of vitamin B₁₂, whereas the other does not. In the latter case, improvement is achieved by administering large (pharmacological) doses of vitamin B₁₂ (1 g per day). Fibroblasts from patients cultured in a medium containing vitamin B₁₂ (at a concentration of 25 pg∙mL^-1) show poor oxidation of [¹⁴C]-propionate. The 5'-deoxyadenosylcobalamin content in cultured cells was only about 10% of that in control cells. When the vitamin B₁₂ concentration in the medium was increased 10,000-fold, both The rate of propionate oxidation and the intracellular concentration of 5'-deoxyadenosylcobalamin approached normal levels. At the same time, no abnormalities in the binding of the coenzyme to the mutase apoenzyme were observed. Thus, the second form of methylmalonic aciduria appears to be caused by an inability to synthesize 5'-deoxyadenosylcobalamin at normal vitamin concentrations.

The above Examples of Inherited Disorders of amino acid catabolism represent relatively well-characterized diseases. A more detailed review of this subject is available elsewhere (Wellner and Meister, 1981).

References

Bremer H. J. et al. Amino acid metabolism: Clinical Chemistry and Diagnosis, Urban and Schwarzenberg. 1981.

Cooper A. J. L. Biochemistry of the Sulfur-Containing Amino Acids, Annu. Rev. Biochem., 1983, 52, 187.

Felig P. Amino acid metabolism in man, Annu. Rev. Biochem., 1975, 44, 933.

Frimter G. W. Aminoacidurias due to disorders of metabolism. (2 parts), N. Engl. J. Med., 1973, 289, 835, 895.

Paxton R., Harris R. A. Isolation of rabbit liver branched chain a-ketoacid dehydrogenase and regulation by phosphorylation, J. Biol. Chem., 1982, 257, 14433.

Paxton R., Harris R.A. Regulation of branched-chain a-ketoacid dehydrogenase kinase, Arch. Biochem Biophys, 1984, 231, 48.

Rosenberg L. E., Scriver C. R. Disorders of amino acid metabolism, Chapter 11. In: Metabolic control and Disease, Bondy P. K., Rosenberg L. E. (eds), Saunders, 1980.

Schwarz V. A Clinical Companion to Biochemical Studies, Freeman, 1978.

Stanbury J. B. et al., (eds), The Metabolic Basis of Inherited Disease, 5th ed., McGraw-Hill, 1983.

Wellner D., Meister A. A survey of inborn errors of amino acid metabolism and transport in man, Annu. Rev. Biochem., 1981,50,911.