Urea Cycle and Ammonium Ion Production

Production of Ammonium Ions and the Urea Cycle

• Amino acids can be converted to carbohydrates when they are not needed for synthesizing nitrogen-containing molecules.
• The nitrogen gets removed and the remaining carbohydrate is converted to pyruvate or a citric acid cycle intermediate for energy production or gluconeogenesis.
• The pathway for amino acid nitrogen disposal converts nitrogen to urea, a nontoxic neutral compound excreted in urine
• Amino acid nitrogen is moved to the urea cycle through transamination, ammonia formation, and urea formation.

Transamination Reactions

• Amino acid nitrogen incorporates into urea after amino acids undergo transamination with α-ketoglutarate to produce glutamate.
• Aminotransferases (transaminases) catalyze reactions that transfer the α-amino group from an amino acid to α-ketoglutarate, producing glutamate.
• About 12 transaminases dispose of nitrogen through glutamate production.
• Aspartate aminotransferase catalyzes N transamination between aspartate and glutamate
• Alanine aminotransferase catalyzes N transamination between alanine and pyruvate
• Pyridoxal phosphate, derived from vitamin B6 (pyridoxine), is necessary for transaminases functioning.

Formation of Ammonia

• Glutamate's oxidative deamination to α-ketoglutarate in the mitochondrial matrix gives off free ammonia.
• Glutamate dehydrogenase catalyzes the reaction, producing either nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate (NADPH)
• The reaction is reversible and incorporates free ammonia into α-ketoglutarate to form glutamate if needed.
• Ammonia that is released in the mitochondrial matrix acts as a precursor for the urea cycle.

Formation of Urea

• The urea cycle starts in the mitochondrial matrix and produces urea in the cytoplasm.
• Carbamoyl phosphate synthetase I (CPS I) joins ammonium ions with carbon dioxide and adenosine triphosphate to produce carbamoyl phosphate.
• Carbamoyl phosphate and ornithine condense to form citrulline via ornithine transcarbamoylase
• Both ornithine and citrulline need specific membrane transport carriers in the mitochondrial membrane.
• Citrulline and aspartic acid condense in the cytoplasm by argininosuccinic acid synthetase, which then forms argininosuccinate
• Argininosuccinate is then cleaved to make fumarate and arginine via argininosuccinase
• Arginase cleaves arginine to release urea and regenerate ornithine.

Anaplerotic Replacement of Aspartate

• An active urea cycle can deplete cytoplasmic aspartate through argininosuccinate formation
• An anaplerotic mechanism compensates by converting fumarate to oxaloacetate (OAA), which can then turn into aspartate.
• These enzymes involved are distinct from those in the mitochondrion.

Urea Cycle Regulation

• During a high-protein meal, catabolism of excess amino acids produces ammonia
• CPS I is allosterically activated by N-acetylglutamate, which is synthesized from acetyl-coenzyme A (CoA) and glutamate, and is stimulated by arginine.
• Sustained elevated ammonia levels activate genes for urea cycle enzymes that occurs during starvation when muscle proteins break down for energy.

Key Points About Production of Ammonium Ions and the Urea Cycle

• Amino acid nitrogen is transferred to the urea cycle as (1) transamination, (2) ammonia formation, and (3) carbamoyl phosphate formation.
• The C skeleton of aspartate is in OAA, and the C skeleton of glutamic acid is in α-ketoglutarate.
• The mitochondrial CPS needs a positive allosteric effector, N-acetylglutamate, for activity, while the cytosolic CPS does not need acetylglutamate and uses glutamine as the N donor for carbamoyl phosphate synthesis.

Amino Acid Degradation

• Transamination produces α-keto acids that enter intermediary metabolism as pyruvate, acetyl-CoA, acetoacetyl-CoA, or citric acid cycle intermediates.
• Ketogenic amino acids convert to acetyl-CoA or acetoacetyl-CoA, with glycogenic amino acids turning into pyruvate or citric acid cycle intermediates.
• Alanine yields pyruvate directly by transamination, whereas cysteine and serine lose side chains before conversion.

Conversion of Aspartate and Asparagine to Oxaloacetate

• Asparaginase removes the amide nitrogen on the asparagine side chain to produce aspartate
• Aspartate converts to OAA via transamination with aspartate aminotransferase.

Branched-Chain Amino Acid Degradation to Succinyl-Coenzyme A and Acetoacetyl-Coenzyme A

• Leucine, isoleucine, and valine are transaminated to form branched-chain α-keto acids, which then undergo oxidative decarboxylation via branched-chain α-ketoacid dehydrogenase multienzyme complexes.
• Valine and isoleucine convert to succinyl-CoA, and leucine converts to acetoacetyl-CoA.

Conversion of Glutamine, Proline, Arginine, and Histidine to α-Ketoglutarate

• Glutamine converts to glutamate via glutaminase, and proline, arginine, and histidine's side chains are modified to produce glutamate.
• Glutamate converts to α-ketoglutarate via glutamate dehydrogenase. Conversion between histidine and glutamate is used to test for folate deficiency.

Conversion of Methionine to Succinyl-Coenzyme A

• Methionine converts to homocysteine in the activated methyl cycle.
• Cystathionine synthase transforms homocysteine to cystathionine, which then converts to propionyl-CoA.
• Propionyl-CoA then converts to succinyl-CoA via methylmalonyl-CoA
• S-adenosyl methionine (SAM) is formed in the activated methyl cycle by transfer of the adenosyl group from adenosine triphosphate to the S of methionine.

Conversion of Phenylalanine and Tyrosine to Fumarate and Acetoacetyl-Coenzyme A

• Phenylalanine and tyrosine degrade to homogentisate and eventually fumarate and acetoacetate.

Amino Acids Degredation Key Points

• Transamination of amino acid nitrogen produces α-keto acids that enter metabolism as pyruvate, acetyl-CoA, acetoacetyl-CoA, and citric acid cycle intermediates.
• Branched-chain amino acids get degraded via ways similar to pyruvate and α-ketoglutarate oxidation.
• Histidine converts to glutamate via FIGLU, which appears in urine with folate deficiencies when given a histidine load.
• SAM forms during activated methyl cycle and serves as methyl donor for the synthesis of hormones, nucleotides, and membrane lipids.

Biosynthesis of Amino Acids and Amino Acid Derivatives

• Nonessential amino acids can be synthesized, whereas essential amino acids must be in the diet.
• Cysteine and tyrosine depends on enough methionine and phenylalanine.

Synthesis of Glutamate, Alanine, and Aspartate

• Glutamate dehydrogenase adds free ammonium ions into α-ketoglutarate to produce glutamate by reversing oxidative deamination.
• Glutamate then acts a nitrogen source via transamination with pyruvate to produce alanine and OAA to produce aspartate.

Synthesis of Glutamine

• Glutamine synthetase produces glutamine from glutamate in an energy-consuming reaction.

Synthesis of Serine and Glycine

• Serine is synthesized via 3-phosphoglycerate converting to 3-phosphopyruvate, which is then transaminated to form 3-phosphoserine.
• Serine forms by phosphate ester removal.
• Glycine forms from serine in a folate-dependent reaction.

Synthesis of Cysteine

• Homocysteine from dietary methionine combines with serine to produce cystathionine
• Cystathionine is then cleaved to produce cysteine, ammonium ion, and α-ketobutyrate, which is then converted to propionyl-CoA.

Synthesis of Catecholamines and Melanin from Phenylalanine and Tyrosine

• Phenylalanine hydroxylase converts phenylalanine to tyrosine.
• Tyrosine is also hydroxylated, which yields 3,4-dihydroxyphenylalanine (DOPA) in neural tissue and the adrenal medulla.
• DOPA then decarboxylates to 3,4-dihydroxyphenylethylamine (dopamine) and further hydroxylated to produce norepinephrine.
• Methylation of DOPA using SAM, produces epinephrine.
• In melanocytes, DOPA oxidizes to dopaquinone, which polymerizes into melanin (the skin pigment).

Synthesis of Serotonin and Melatonin

• Tryptophan hydroxylase converts tryptophan to 5-hydroxytryptophan and converted to serotonin (5-hydroxytryptamine)
• Serotonin synthesis is located in the hypothalamus and brainstem, pineal gland, and chromaffin cells of the gut.
• Melatonin is produced from serotonin in the pineal gland during the dark phase of the light/dark cycle to regulate the sleep/wake cycle

Neurotransmitters

• γ-Aminobutyrate (GABA) is synthesized by decarboxylation of glutamate. GABA is inhibitory, likewise the monocarboxylic amino acids glycine, β-alanine, and taurine. Dicarboxylic amino acids glutamate and aspartate are excitatory.

Heme Synthesis

• The rate-limiting step in heme synthesis is the condensation of succinyl-CoA and glycine to form δ-aminolevulinic acid (ALA).
• ALA synthetase catalyzes messenger RNA translation that occurs in heme, feedbacking to inhibit its own synthesis.
• ALA dehydratase catalyzes 2 molecules of ALA to form porphobilinogen
• Defective ALA dehydratase caused by lead poisoning results in accumulating ALA
• The porphyrin ring then turns into coproporphyrinogen III in the cytoplasm followed by transportation to mitochondrion for conversion to protoporphyrinogen IX
• Ferrous atom then gets added to protoporphyrin IX by ferrochelatase.

Heme Degradation

• Heme oxygenase opens the tetrapyrrole ring, which creates biliverdin (verd = green) and carbon monoxide.
• This uses NADPH and molecular O2. Next, biliverdin reductase results in bilirubin via an NADPH-requiring reaction.
• Bilirubin, a hydrophobic compound, is bound by albumin and transported to the liver, where it is conjugated with two molecules of glucuronic acid to produce bilirubin diglucuronide, which is excreted in bile.

Bilirubin Metabolism in the Gut

• Gut flora hydrolyzes bilirubin diglucuronide and reduces free bilirubin to the colorless urobilinogen
• Urobilinogen degrades to stercobilin, giving feces its characteristic brown color.
• Some urobilinogen reabsorbs from the gut and is removed from the circulation in urine as urobilin, coloring urine like amber.

Phenylketonuria

• Phenylketonuria (PKU) presents with elevated blood phenylalanine that leads to mental retardation beginning in utero.

Primary Phenylketonuria

• Deficient enzyme in the primary form of PKU, PKU I, is phenylalanine hydroxylase
• Accumulation of phenylalanine causes increases products by a minor pathway which leads to neurotoxic results like phenylpyruvate, phenylacetic acid, and phenyllactic acid
• A phenylalanine-restricted diet can prevent neurologic damage until 6 years of age

Secondary Phenylketonuria

• Deficient dihydrobiopterin reductase causes secondary form, PKU II
• Restricted blood levels via diet are found, but neurologic damage remains unaffected

Alcaptonuria

• Homogentisate accumulates and causes urine is oxidized into a dark color

Methylmalonic Acidemia

• A deficiency in methylmalonyl-CoA mutase functions in the conversion of methionine, isoleucine, and valine to succinyl-CoA is presented
• Results in affected newborns with recurrent vomiting, hepatomegaly, and developmental retardation

Maple Syrup Urine Disease

• A deficiency of branched in branched-chain α-keto acid dehydrogenase enzyme causes maple syrup urine disease or branched-chain ketonuria.
• Affected infants have significant vomiting and severe mental defects.
• Therapy includes branched-chain amino acids restrictions

Urea Cycle Disorders

• All defects in the urea cycle result in interference with ammonia excretion which leads to ammonia toxicity (hyperammonemia) during CPS or OTC defects Citrullinemia and argininosuccinic aciduria are treated with arginine.
• Sodium benzoate and phenylacetate help by excreting urine.
• Causes adducts with glycine (hippuric acid is benzoylglycine) and glutamine that consumes nitrogen to replace glycine and glutamine

Porphyrias

• Results from deficiency of a biosynthetic pathway enzyme
• Usually accompanied by photosensitivity Acute intermittent porphyria: Buildup of porphobilinogen and ALA Porphyria cutanea tarda: From building up porphyrins that demonstrates photosensitivity if deficient in uroporphyrinogen decarboxylase