Biochemistry - 42 - Amino Acid Synthesis and Degradation 2023 PDF

Amino Acid Synthesis and Degradation Lecture 42 Reference: Lieberman and Peet, Chapter 37 Bluefield University – VCOM Campus MABS Program Biochemistry Jim Mahaney, PhD See Notes Learning Objectives a. Recall the synthesis of the non-essential amino acids often starts with intermediates of glycolys...

Amino Acid Synthesis and Degradation Lecture 42 Reference: Lieberman and Peet, Chapter 37 Bluefield University – VCOM Campus MABS Program Biochemistry Jim Mahaney, PhD See Notes Learning Objectives a. Recall the synthesis of the non-essential amino acids often starts with intermediates of glycolysis or the TCA (tricarboxylic acid) cycle. b. Relate the enzyme deficiencies that cause cystathioninurea and homocystinuria types I, II and III. c. Recall that tyrosine is synthesized from phenylalanine and identify the enzyme that does this reaction. d. Compare and contrast classical versus malignant phenylketonuria, including the enzymes that are deficient in each case and the metabolic consequences of each type of PKU. e. Recall that catabolism of the carbon chains of amino acids lead to glycolytic intermediates, pyruvate, acetyl CoA or acetoacetate. f. Relate the terms glucogenic and ketogenic amino acids. g. Recall the enzyme deficiency that causes histidinemia. h. Recall the major deficiencies in tyrosine metabolism, including alcaptonuria and tyrosinemia. Identify which enzyme is deficient by which metabolic consequence is present. i. Recall methylmalonyl CoA mutase as a vitamin B12-dependent enzyme that brings amino acid carbons into the TCA cycle at succinyl CoA. j. Identify the three branched chain amino acids (BCAA) and recall the enzyme deficiency in BCAA degradation that causes maple syrup urine disease. 2 Objectives A and E Overview • Humans can synthesize only 11 of the 20 amino acids required for protein synthesis. (“non-essential”) • The other 9 amino acids are considered “essential” and must be obtained from the diet. • The non-essential amino acids can be synthesized from glycolytic intermediates, TCA cycle intermediates, or from existing amino acids. • When amino acids are degraded, the nitrogen is converted to urea, and the carbon skeletons are classified as glucogenic (precursor for glucose) or ketogenic (precursor of ketone bodies) • Defects in amino acid degradation pathways can lead to disease. Non-essential Alanine Arginine Aspartate Asparagine Cysteine Glutamate Glutamine Glycine Proline Serine Tyrosine Essential Histidine Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine 3 Biosynthesis of Non-Essential Amino Acids Objective A • The 11 non-essential amino acids are synthesized from intermediates of metabolism • NOTE: Cys and Tyr require an essential amino acid • Amino acids from glycolytic intermediates • • • • Glycine Serine Cysteine Alanine • Amino acids from TCA cycle intermediates • Asparagine • • • • Aspartate Glutamate Glutamine Proline • Arginine 4 Synthesis from α-Keto Acids (Ala, Asp, Glu) • Alanine, aspartate, and glutamate are synthesized by transfer of an amino group to the α-keto acids pyruvate, oxaloacetate, and αketoglutarate respectively • These transamination reactions are the most direct of the biosynthetic pathways • NOTE: Glutamate is unusual in that in can also be synthesized by the reverse of oxidative deamination, catalyzed by glutamate dehydrogenase when NH3 is high 5 Synthesis by Amidation (Gln, Asn) Glutamine • Produced from glutamate by glutamine synthetase (adds a NH3 to the γ-carboxyl group of the side chain) • The reaction is driven by the hydrolysis of ATP • Glutamine is reconverted to glutamate by glutaminase (*important in kidney) • Major mechanism for the transport of ammonia in a non-toxic form Asparagine • Asparagine is formed from aspartate by asparagine synthetase in a reaction in which glutamine provides the nitrogen for the amide group. (*differs from synthesis of glutamine) • The reaction is driven by the hydrolysis of ATP • Certain types of tumor cells (leukemic) require asparagine for growth. • Asparaginase has been used as an antitumor agent 6 Synthesis of Proline and Arginine Proline • Glutamate is 1st phosphorylated an then converted to glutamate semialdehyde (reduction of side chain) • The semialdehyde group spontaneously cyclizes and reduction of cyclic compound yields proline • Proline is converted back to glutamate semialdehyde Arginine [not shown] • Arginine is synthesized from glutamate via glutamate semialdehyde • Quantity of arginine only adequate for adults and insufficient to support growth. • During periods of growth, arginine is an essential amino acid • Arginine is cleaved by arginase to form urea and ornithine 7 Synthesis of Serine • All derived from intermediates of glycolysis • Serine is synthesized from 3-phosphoglycerate and produces glycine and cysteine Serine • Serine arises from 3-phosphoglycerate (an intermediate of glycolysis) • 3-phosphoglycerate is first oxidized to 3-phosphopyruvate and then transaminated to 3-phosphoserine • Serine if formed by hydrolysis of the phosphate ester • High serine levels repress 3-phosphoglycerate dehydrogenase and phosphoserine phosphatase (negative feedback) 8 Synthesis of Glycine and Cysteine Glycine Major pathway • Glycine is synthesized from serine by removal of a hydroxymethyl group by serine hydroxymethyltransferase (major pathway) • Tetrahydrofolate (THF) is the one-carbon acceptor • In a minor pathway, glycine can be synthesized from threonine by removal of acetaldehyde by threonine aldolase (PLP required) Cysteine • Carbon skeleton and nitrogen provided by serine, and the sulfur is provided by methionine (essential). • Ser reacts with homocysteine (from Met) to form cystathionine (cystathionine βsynthase) • Cleavage of cystathionine by cystathionase produces cysteine and α-ketobutyrate • Cys regulates its own production to adjust for dietary supply by inhibiting cystathionine β-synthase • Cys becomes essential if methionine is low in the diet. Adequate cysteine in the diet “spares” met (i.e., don’t have to use precious met make cys). 9 Cystathioninuria Objective B Cystathioninuria: the presence of cystathionine in the urine. • Accumulation of cystathionine and its metabolites is due to a rare deficiency in cystathionase [Recall: cystathionase and PLP convert cystathionine to cysteine and α-ketobutyrate] • Common in premature infants. (As they mature enzyme levels rise and urine cystathionine levels decline) • Adults: Caused by genetic cystathionase deficiency or dietary deficiency of pyridoxine (vitamin B6) • No characteristic clinical abnormalities-probably a benign disorder 10 Homocystinuria Type I Objective B Homocystinuria: Abnormal elevation of homocysteine and its metabolites in the blood and urine. • Homocystinuria is an autosomal recessively inherited defect in the transsulfuration pathway (Type I) or methylation pathway (Type II and III). • Type I is due to a deficiency in cystathionine β-synthase or dietary deficiency of Vitamin B6 • Renal tubular reabsorption of Methionine is efficient, so it may not appear in urine. But homocysteine is less efficiently reabsorbed, thus large amounts excreted in urine daily in homocystinuria. • Elevations promote oxidative damage, inflammation, and endothelial dysfunction. ~ALSO~ risk factor for vascular disease 11 Homocystinuria Types II and III Objective B Homocystinuria: Abnormal elevation of homocysteine and its metabolites in the blood and urine. Type III Type II Homocystinuria • Deficiency in the synthesis of methyl- B12 (CH3-B12 or cobalamin) Type III Homocystinuria Type II • Deficiency in the synthesis of methyltetrahydrofolate (N5- CH3-FH4) BOTH associated with elevated plasma homocysteine but not methionine **Methionine can’t be regenerated ** 12 Synthesis of Tyrosine Objective C and D • Phenylalanine is hydroxylated to form tyrosine by phenylalanine hydroxylase (PAH) • The reaction requires molecular oxygen and the coenzyme tetrahydrobiopterin (BH4) • BH4 (redox cofactor) is oxidized to BH2 and MUST be reduced back to BH4 for PAH to continue forming tyrosine • BH4 is not synthesized from a vitamin; it can be generated from GTP • Phenylketonuria (PKU) results from deficiencies of PAH (classic form); dihydropteridine reductase, or enzymes in the biosynthetic pathway for BH4 (malignant form) 13 Malignant Phenylketonuria (PKU) Objective D Definition: Subset (~3%) of hyperphenylalanemia patients that show an appropriate reduction in plasma Phe levels with dietary restriction; however progressive neurological symptoms and seizures still develop with death occurring within 2 years of life. • Characteristics: – Normal PAH activity – Deficiency in dihydropteridine reductase (DHPR) – enzyme required for regeneration of BH4, a cofactor of PAH causing an indirect increase in Phe levels – Less frequently, DHPR activity is normal but a defect in the biosynthesis of BH4 exists • Dietary therapy corrects hyperphenylalanemia • BH4 is also a cofactor for 2 other hydroxylation's required in the synthesis of neurotransmitters in the brain [TrpàSerotonin and TyràCatecholamines] – It has been suggested that the resulting deficit in the CNS neurotransmitter activity is at least in part responsible for the neurologic manifestations and death of these patients. 14 Amino Acid Degradation Objective E, F • As amino acids are degraded, their carbons are converted to: • CO2 • Compounds that produce glucose in the liver ( pyruvate, α-KG, succinyl Co-A, fumarate, oxaloacetate) • Ketone bodies or their precursors (Acetyl CoA, acetoacetate) Glucogenic Ketogenic • Amino acids can be classified as glucogenic, ketogenic, or both, based on which of the 7 intermediates are produced during their catabolism • Glucogenic amino acids – carbon skeletons can be converted to pyruvate or one of the intermediates of the TCA cycle • These intermediates are substrates for gluconeogenesis and therefore, can give rise to the net synthesis of glucose in liver and kidney • Ketogenic amino acids – carbon skeletons can be converted to acetoacetate or one of its precursors (acetyl-CoA or acetoacetylCoA) • Acetoacetate is one of the ketone bodies, which also include 3-hydroxybutyrate and acetone • Lysine and leucine are the only exclusively ketogenic amino acids 15 Essential Nonessential Glucogenic vs. Ketogenic Amino Acids Glucogenic Glucogenic and Ketogenic Alanine Arginine Asparagine Aspartate Cysteine Glutamate Glutamine Glycine Proline Serine Tyrosine Histidine Methionine Valine Isoleucine Phenylalanine Threonine Tryptophan Objective F Ketogenic Leucine Lysine 16 Amino Acids that Form Acetyl CoA or Acetoacetyl CoA Ketogenic Degradation Leucine, isoleucine, lysine, threonine, and tryptophan form acetyl CoA or acetoacetyl CoA directly Leucine (strictly ketogenic) • produces hydroxymethylglutaryl CoA (HMG-CoA), which is cleaved to form acetyl-CoA and acetoacetate. • Primary metabolism occurs in muscle. Isoleucine (ketogenic and glucogenic) • Metabolism yields acetyl CoA and propionyl CoA Lysine (strictly ketogenic) • Cannot be directly transaminated at either of its two amino groups • Degraded by a complex pathway which generates NADH and FADH2 for energy • Ultimately generates acetyl-CoA Threonine (ketogenic and glucogenic) • A minor pathway in threonine degradation by threonine aldolase produces glycine and acetyl-CoA Tryptophan (ketogenic and glucogenic) • Catabolism yields alanine and acetoacetyl CoA 17 Amino Acids that Form Pyruvate Glucogenic Degradation Alanine • Alanine loses its amino group by transamination to form pyruvate Serine • Serine can be converted to glycine and N5,N10methylene-FH4. Serine can also be converted to pyruvate by serine dehydratase Glycine • Glycine can be converted to serine by the reversible addition of a methylene group from N5,N10-methyleneFH4 or oxidized to CO2 and NH4+ 18 Amino Acids that Form Pyruvate Glucogenic Degradation Glycine (cont.) • Glycine can be deaminated to glyoxylate, which can be oxidized to oxalate or transaminated to glycine. Lack of transaminase leads to primary oxaluria type I (PH1) [renal failure] Oxalate precipitates in the kidney leading to kidney stones • Deficiency of the transaminase in liver peroxisomes causes overproduction of oxalate, the formation of oxalate stones, and kidney damage (primary oxaluria type 1 or PH1) 19 Amino Acids that Form Pyruvate Glucogenic Degradation Cysteine • Cysteine undergoes desulfuration to yield pyruvate • The sulfate released can be used to synthesize 3’phosphoadenosine-5’-phosphosulfate (PAPS), an activated sulfur donor to a variety of acceptors • Cysteine can be oxidized to its disulfide derivative, cystine. Threonine • Threonine is converted to pyruvate in most organisms but it a minor pathway (at best) in humans 20 Amino Acids That Form Oxaloacetate Glucogenic Degradation • Asparagine is hydrolyzed by asparaginase, liberating ammonium (NH4+) and aspartate • Aspartate loses its amino group by transamination to form oxaloacetate • Some rapidly dividing leukemic cells are unable to synthesize sufficient asparagine to support their growth • This makes asparagine an essential amino acid for these cells, which therefore requires asparagine from the blood. • Asparaginase, which hydrolyzes asparagine to aspartate can be administered systemically to treat leukemic patients. • Asparaginase lowers the level of asparagine in the plasma, thereby depriving cancer cells of a required nutrient. 21 Amino Acids that Form α-Ketoglutarate via Glutamate Glucogenic Degradation Glutamine • Glutamine is hydrolyzed to glutamate and ammonium by the enzyme glutaminase [see slide 7] • Glutamate is converted to α-ketoglutarate by transamination or through oxidative deamination by glutamate dehydrogenase [review lecture 47] Proline • Proline is oxidized to glutamate. Glutamate is transaminated or oxidatively deaminated to form α-ketoglutarate. Arginine • Arginine is hydrolyzed by arginase to produce ornithine (and urea). [Review lecture 47] • Ornithine is subsequently converted to α-ketoglutarate, with glutamate semialdehyde as an intermediate 22 Amino Acids that Form α-Ketoglutarate via Glutamate Glucogenic Degradation • Histidine is an essential amino acid and cannot be synthesized by humans • 5 of its carbons form glutamate when degraded. • Histidine is oxidatively deaminated by histidase to urocanic acid which subsequently forms N-formiminoglutamate (FIGLU) • The subsequent reactions transfer 1 carbon of FIGLU to the tetrahydrofolate (FH4) pool and release NH4+ and glutamate. • Glutamate is transaminated or oxidatively deaminated to form αketoglutarate. Histidinemia: rare genetic disorder caused by histidase deficiency. Causes elevated histidine level in blood. Generally considered benign. Objective G 23 Amino Acids that Form Fumarate Objective H Glucogenic Degradation Phenylalanine and tyrosine • Hydroxylation of phenylalanine produces tyrosine in a reaction catalyzed by BH4 requiring phenylalanine hydroxylase (PAH), initiating the catabolism of Phe • Tyrosine undergoes oxidative degradation in a pathway that produces fumarate and acetoacetate • Inherited deficiencies in the enzymes of phenylalanine and tyrosine metabolism lead to the diseases: • PKU [see slides 15-17] • Tyrosinemia • Alkaptonuria 24 Tyrosinemia Objective H • Transient tyrosinemia is frequently observed in newborn infants (especially premature) – Condition appears to be benign – Dietary restriction of protein returns plasma Tyr to normal. – Biochemical defect is most likely a low level immaturity of 4-hydroxyphenylpyruvate dioxygenase • Tyrosinemia II: deficiency in tyrosine aminotransferase (TAT) [Tyr à p-Hydroxyphenylpyruvate] – Tyrosine accumulates in the urine. – Leads to eye and skin lesions, neurological symptoms. – Treated by restricting Tyr and Phe intake • Tyrosinemia I (Tyrosinosis): deficiency in fumarylacetoacetate hydrolase. [last step] – – – – Fumarylacetoacetate and its metabolites, particularly succinyl-acetone accumulate in the urine. Acute form associated with liver failure, renal tubular acidosis and characteristic cabbage-like odor. Treatment includes dietary restriction of Phe and Tyr Without treatment death within first year of life. 25 Alkaptonuria Objective H • CAUSE: Inherited genetic deficiency of homogentisate oxidase resulting in an accumulation of homogentistic acid (HGA) in the skin and other tissues. The acid leaves the body through the urine turning brownish-black when it mixes with air. • SYMPTOMS: Infant urine may darken, arthritis (especially of spine), darkening of ear, dark spots on sclera and cornea. • TREATMENT: Some patients benefit from high-dose of vitamin C. This has shown to decrease the build-up of brown pigment in cartilage, may slow arthritis. • PROGNOSIS: Life expectancy is unaffected. All patients will experience chronic joint pain. • POSSIBLE COMPLICATIONS: Arthritis in ~50% of older adults, heart valve replacement, coronary heart disease, and kidney stones. • PREVENTION: None known 26 Amino Acids that Form Succinyl-CoA Objective I Glucogenic Degradation Methionine • Methionine condenses with ATP, forming S-adenosylmethionine (SAM) • The methyl group attached to SAM is “activated” and can be transferred by methyltransferases to a variety of acceptor molecules [see lecture 51], the reaction product is S-adenosylhomocysteine (SAH) • SAH is hydrolyzed to homocysteine and adenosine • Homocysteine has 2 fates: Methylmalonyl CoA mutase • If there is a deficiency in Methionine, homocysteine is remethylated to methionine [requires FH4 and B12] • If methionine stores are adequate, homocysteine may enter the transsulfuration pathway, where it is converted to cysteine. Note: Methylmalonyl CoA mutase converts methylmalonyl CoA to succinyl CoA (TCA Cycle intermediate). 27 Objective I Amino acids that Form Succinyl-CoA: Continued Valine and Isoleucine • Branched-chain amino acids (BCAAs) that generate propionyl-CoA, which is converted to methylmalonyl CoA and then succinyl-CoA by biotin and B12 requiring reactions Methylmalonyl CoA mutase Threonine • Primarily degraded by a PLP-requiring dehydratase to ammonia and α-ketobutyrate. • Oxidative decarboxylation to form propionyl-CoA • Propionyl-CoA is converted to succinyl-CoA 28 Methylmalonyl CoA Mutase (MCM) Deficiency Objective I • Branched-chain amino acids and methionine degradation all lead into SuccinylCoA. • MCM catalyzes the isomerization of methylmalonyl-CoA to succinyl-CoA • MCM requires a vitamin B12-derived prosthetic group to function and is involved in key metabolic pathways • Genetic deficiency in this mutase gene causes failure to thrive, vomiting, dehydration, developmental delay, and seizures • Can also lead to methylmalonic acidemia 29 Catabolism of Branched Chain Amino Acids Glucogenic Degradation • The BCAAs isoleucine, leucine, and valine are essential amino acids that are primarily metabolized by peripheral tissues (particularly muscle) rather than the liver [universal fuels] • Degradative pathway has 2 major functions: (1) energy (2) provide precursors to replenish TCA cycle intermediates • 1st step is transamination by B6 requiring branched-chain amino acid aminotransferase forming an α-ketoacid • 2nd step: α-keto analogs undergo oxidative decarboxylation by the branched chain α-keto acid dehydrogenase (BCKD) complex • Subsequent degradation steps are analogous to those for β-oxidation of fatty acids, so NADH and FAD(2H) are generated for energy production. • NOTE: Errors in the dehydrogenases of the second step leads to Maple Syrup Urine Disease (very sweet smelling urine – now diagnosed by genetic tests and specific blood tests). Objective J 30 Sample Question When an amino acid is catabolized for energy, what happens to the amino group? A. B. C. D. E. The amino group is removed by transamination The amino group is added to the urea molecule The amino group is transferred onto a glycolytic or TCA cycle intermediate The amino group is used as energy to drive glucose formation via gluconeogenesis The amino group remains on the amino acid molecule 31 Sample Question Which of the following enzyme deficiencies would prevent the synthesis of tyrosine from phenylalanine? A. phenylalanine hydroxylase B. phenylalanine oxidase C. phenylalanine peroxidase D. phenylalanine reductase E. tetrahydrofolate reductase 32 Thank You! 33