BIOCHEMISTRY - Textbook - Ostapchenko L. I. - 2012

Chapter 6. AMINO ACID METABOLISM AND FUNCTIONS. PROTEIN BIOSYNTHESIS

6.8. Metabolism of Individual Amino Acids

6.8.3. Phenylalanine and Tyrosine Metabolism

Phenylalanine is an essential amino acid because its benzene ring cannot be synthesized in animal Cells. Tyrosine is a conditionally essential amino acid, as it is produced from phenylalanine. The content of these Amino Acids in dietary Proteins, including plant-based ones, is relatively low. Phenylalanine and tyrosine are utilized for the synthesis of numerous biologically active compounds. The METABOLISM of these amino acids varies across different tissues (Fig. 6.28).

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Fig. 6.28. Pathways of phenylalanine and tyrosine conversion in various tissues:

H₄BH₄ - tetrahydrobiopterin; H₂BH₄ - dihydrobiopterin;

PLP - Pyridoxal phosphate; SAM - S-adenosylmethionine

The bulk of phenylalanine is utilized via two main pathways:

✵ incorporation into proteins;

✵ conversion into tyrosine.

The conversion of phenylalanine into tyrosine is primarily necessary to eliminate excess phenylalanine, as high concentrations of it are cytotoxic. The synthesis of tyrosine itself is not critically important, since a deficiency of this amino acid in cells is virtually never observed.

The main metabolic pathway of phenylalanine begins with its hydroxylation (Fig. 6.29), yielding tyrosine. This reaction is catalyzed by a specific monooxygenase, phenylalanine hydroxylase, which requires tetrahydrobiopterin (H₄BH₄) as a coenzyme. Enzyme activity also depends on the presence of Fe²⁺. The reaction is irreversible: H₄BH₄ is oxidized during the process to dihydrobiopterin (H₂BH₄). The latter is regenerated by dihydropteridine reductase using NADPH + H+.

Tyrosine metabolism is significantly more complex than that of phenylalanine. Aside from its incorporation into proteins, tyrosine serves in various tissues as a precursor for compounds such as catecholamines, thyroxine, and Melanins, and is catabolized to CO₂ and H₂O.

In the Liver, tyrosine Catabolism proceeds down to its ultimate end products. This specific catabolic pathway comprises several enzymatic reactions and concludes with The formation of fumarate and acetoacetate:

1) Tyrosine Transamination with α-ketoglutarate is catalyzed by tyrosine aminotransferase (with PLP as a coenzyme), an inducible enzyme found in the mammalian liver. This reaction yields p-hydroxyphenylpyruvate.

2) The oxidation of p-hydroxyphenylpyruvate to homogentisic acid involves decarboxylation, aromatic ring hydroxylation, and side-chain migration. This reaction is catalyzed by p-hydroxyphenylpyruvate dioxygenase, which requires Vitamin C and Fe^{2 +} as Cofactors.

3) The conversion of homogentisic acid to fumarylacetoacetate is accompanied by the Cleavage of the aromatic ring. This reaction is catalyzed by homogentisate dioxygenase, which contains Fe²⁺ as a coenzyme.

The Metabolism of Phenylalanine and tyrosine involves numerous hydroxylation Reactions Catalyzed by oxygenases. Oxygenase Enzymes (hydroxylases) utilize an O₂ molecule and a hydrogen donor coenzyme (most commonly H₄BH₄). These enzymes require cofactors such as Fe²⁺ or heme (and Cu⁺ for certain types), while many also require vitamin C. Oxygenases are divided into two groups:

✵ Monooxygenases — incorporate one atom of O₂ into the reaction product, while the second is used to form H₂O;

✵ Dioxygenases — incorporate both atoms of O₂ into the reaction product.

Virtually all aromatic ring cleavage processes in biological systems are catalyzed by dioxygenases, a subclass of enzymes discovered by the Japanese biochemist Osamu Hayaishi. Ring opening of the benzene ring yields maleylacetoacetate, which is subsequently converted into fumarylacetoacetate via cis- and trans-isomerization.

4) The Hydrolysis of fumarylacetoacetate by fumarylacetoacetate hydrolase yields fumarate and acetoacetate. Fumarate can be oxidized to CO₂ and H₂O or used in Gluconeogenesis. Acetoacetate is a ketone body that is oxidized to final products with the release of energy.

In pigment cells (melanocytes), tyrosine serves as a precursor for dark pigments called melanins. Two main types predominate: eumelanins and pheomelanins. Eumelanins (black and brown pigments) are insoluble, high-molecular-weight heteropolymers of 5,6-dihydroxyindole and some of its precursors. Pheomelanins are yellow or reddish-brown polymers that are soluble in dilute alkalis. They are found primarily in Hair. Melanins are also present in the retina of the eye. Skin color depends on the distribution of melanocytes and The amount of various melanin types they contain.

Melanin synthesis is a complex, multi-step, and branched process (for a brief outline of the synthesis pathway, see Fig. 6.28). The first reaction—the conversion of tyrosine to DOPA—is catalyzed by tyrosinase, which utilizes Cu⁺ ions as a cofactor (Scheme A):

The Thyroid Gland synthesizes and secretes iodothyronine Hormones: thyroxine (tetraiodothyronine) and triiodothyronine. These are iodinated tyrosine residues that enter the thyroid follicular cells through the basement membrane (Scheme B):

In The adrenal medulla and Nervous Tissue, tyrosine acts as a precursor for catecholamines (dopamine, norepinephrine, and epinephrine):

During the formation of catecholamines in nervous tissue and the Adrenal Glands, and melanin in melanocytes, dihydroxyphenylalanine (DOPA) serves as an intermediate product. However, the hydroxylation of tyrosine in different Cell types is catalyzed by distinct enzymes:

✵ tyrosinase in melanocytes is a Cu⁺-dependent enzyme;

✵ tyrosine hydroxylase (1) in the adrenal glands and catecholaminergic Neurons does not require copper ions. It is an Fe²⁺-dependent enzyme that, similarly to phenylalanine hydroxylase, uses H₄B as a coenzyme. The Physiological Role of tyrosine hydroxylase is exceptionally significant, as this enzyme is regulatory and determines The rate of catecholamine synthesis. The activity of tyrosine hydroxylase changes significantly due to, first, Allosteric Regulation (with norepinephrine acting as an inhibitor); second, phosphorylation/dephosphorylation: phosphorylation mediated by protein kinase A lowers the K_m for the coenzyme H₄B and the affinity of the enzyme for norepinephrine, resulting in the activation of tyrosine hydroxylase. The amount of the enzyme is regulated at the transcriptional level;

✵ DOPA decarboxylase (2) (with PLP as a coenzyme) catalyzes the formation of dopamine, which is then converted into norepinephrine by dopamine hydroxylase (3) (a monooxygenase). This Enzymatic Function requires Cu⁺ ions, vitamin C, and tetrahydrobiopterin;

✵ in the adrenal medulla, phenylethanolamine N-methyltransferase (4) catalyzes the methylation of norepinephrine to produce epinephrine, with SAM serving as the methyl group donor.

Dopamine and norepinephrine function as Neurotransmitters in synaptic transmission, whereas epinephrine is a broad-spectrum hormone that regulates Energy Metabolism. One of the key Functions of catecholamines is the Regulation of The Cardiovascular system.

Several inherited disorders are known to be associated with defects in the enzymes of Phenylalanine and Tyrosine Metabolism across various tissues. In the liver of healthy individuals, a small fraction of phenylalanine (~10%) is converted into phenyllactate and phenylacetylglutamine (Fig. 6.30).

This pathway of phenylalanine catabolism becomes predominant when the primary pathway—its conversion to tyrosine catalyzed by phenylalanine hydroxylase—is impaired. Such a disruption leads to hyperphenylalaninemia and an increased level of alternative pathway metabolites in the Blood and urine: phenylpyruvate, phenylacetate, phenyllactate, and phenylacetylglutamine. A defect in phenylalanine hydroxylase is the underlying cause of phenylketonuria (PKU), which exists in two forms:

✵ classical PKU is an inherited disorder caused by Mutations in the phenylalanine hydroxylase Gene that lead to reduced enzyme activity or its complete inactivation. As a result, blood phenylalanine concentration increases 20-30 fold (normal range is 1.0-2.0 mg/dL), and urinary concentration increases 100-300 fold compared to normal levels (30 mg/dL). The concentrations of phenylpyruvate and phenyllactate in the urine reach 300-600 mg/dL, whereas they are completely absent under normal conditions.

Fig. 6.30. Alternative pathways of phenylalanine catabolism.

Due to the phenylalanine hydroxylase defect, the accumulated phenylalanine undergoes transamination with α-ketoglutarate. The resulting phenylpyruvate is converted into either phenyllactate or phenylacetylglutamine, which accumulate in the blood and are excreted in the urine.

These compounds are toxic to Brain cells.

The most severe manifestations of PKU include impaired mental and physical development, abnormal pigmentation, and seizure disorders. If left untreated, patients typically do not survive past the age of 30. The incidence of the disease is 1 in 10,000, and it is inherited in an autosomal recessive manner.

The severe manifestations of PKU are associated with the Toxic effects of high concentrations of phenylalanine, phenylpyruvate, and phenyllactate on brain cells. Elevated levels of phenylalanine restrict The transport of tyrosine and Tryptophan across the blood-brain barrier and inhibit the synthesis of neurotransmitters (dopamine, norepinephrine, serotonin).

✵ variant PKU (coenzyme-dependent hyperphenylalaninemia) arises from mutations in genes that control H₄B metabolism. The clinical manifestations are nearly identical to those of classical PKU. The incidence is 1-2 cases per 1 million newborns.

H₄B is required for the hydroxylation reactions not only of phenylalanine but also of tyrosine and tryptophan; therefore, a deficiency of this coenzyme disrupts the metabolism of all Three amino acids, including neurotransmitter synthesis. The condition is characterized by severe neurological disorders and early mortality ("malignant PKU").

Progressive impairment of mental and physical development in children with PKU can be prevented by a very low-phenylalanine or phenylalanine-free diet. If initiated right after birth, brain damage can be averted entirely. It is generally believed that dietary restrictions can be relaxed after the age of 10 (following the completion of brain myelination processes), but many contemporary pediatricians advocate for a lifelong diet.

PKU is diagnosed using Qualitative and quantitative Methods to detect pathological metabolites in urine, as well as by measuring phenylalanine concentrations in blood and urine. The defective gene responsible for phenylketonuria can be identified in phenotypically normal heterozygous carriers using a phenylalanine tolerance test. In this Procedure, the subject is administered ~10 g of phenylalanine in solution on an empty Stomach, followed by blood samples taken at hourly intervals to determine tyrosine levels. Normally, the blood tyrosine concentration after a phenylalanine load is significantly higher than that in heterozygous carriers of the phenylketonuria gene. This test is utilized in Genetic Counseling to assess the risk of having an affected child. Screening protocols have been developed to identify newborn infants with PKU, achieving a test sensitivity of nearly 100 %.

Today, mutations in the gene responsible for PKU can be diagnosed using DNA-based methods (such as restriction fragment length polymorphism analysis and PCR).

Certain disorders of hepatic tyrosine catabolism lead to tyrosinemia and tyrosinuria. Three types of tyrosinemia are distinguished.

Type I tyrosinemia (tyrosinosis). The condition is likely caused by a deficiency of fumarylacetoacetate hydrolase, the enzyme that catalyzes The breakdown of fumarylacetoacetate into fumarate and acetoacetate (see Fig. 6.28). The accumulation of these metabolites inhibits several enzymes and Amino Acid Transport systems. The Pathophysiology of this disorder is rather complex. The acute form of tyrosinosis typically manifests in newborns, presenting clinically with diarrhea, vomiting, and developmental delays. Without Treatment, affected infants die at 6-8 months of age due to progressive Liver failure. The chronic form features similar but less pronounced symptoms, with mortality occurring by age 10. Blood tyrosine levels in patients are several times higher than normal. Treatment involves a diet restricted in both tyrosine and phenylalanine.

Type II tyrosinemia (Richner-Hanhart syndrome). The underlying cause is a deficiency of the enzyme tyrosine aminotransferase. Blood tyrosine concentrations are elevated in patients. The disease is characterized by ocular and cutaneous lesions, moderate mental retardation, and impaired motor coordination.

Neonatal tyrosinemia (transient). This condition arises from reduced activity of p-hydroxyphenylpyruvate dioxygenase, the enzyme that converts p-hydroxyphenylpyruvate into homogentisic acid (see Fig. 6.28). Consequently, the blood concentrations of p-hydroxyphenylacetate, tyrosine, and phenylalanine are elevated. Treatment involves a low-protein diet and vitamin C supplementation.

Alkaptonuria ("black urine disease"). The cause of the disorder is a deficiency of homogentisic acid dioxygenase (Fig. 6.28). This condition is characterized by the urinary excretion of large amounts of homogentisic acid, which oxidizes upon exposure to air, forming dark pigments known as alkaptons. This metabolic anomaly was described as early as the 16th century, and the disease itself was characterized in 1859. In addition to the darkening of urine upon standing, clinical manifestations include Connective Tissue pigmentation (ochronosis) and Arthritis. The incidence is 2–5 cases per 1 million newborns. The disease is inherited in an autosomal recessive manner. Diagnostic methods for identifying heterozygous carriers of the defective gene have not yet been developed.

Albinism. The metabolic defect is a congenital deficiency of tyrosinase, the enzyme that catalyzes the conversion of tyrosine to DOPA in melanocytes. Consequently, melanin synthesis is impaired.

Clinically, albinism (from Latin *albus* — white) manifests as a complete lack of skin and hair pigmentation. Affected individuals often suffer from reduced visual acuity and photophobia. Prolonged exposure to direct sunlight puts these patients at a high risk for skin Cancer. The incidence of albinism is approximately 1 : 20,000.

Impaired catecholamine synthesis (Fig. 6.28) can lead to various neuropsychiatric disorders, with pathological abnormalities arising from both decreased and increased neurotransmitter levels.

Parkinson's disease. This condition develops as a result of dopamine depletion in the substantia nigra of the brain. It is one of the most common neurological disorders (affecting approximately 1 : 200 individuals over the age of 50). This pathology is marked by decreased activity of tyrosine hydroxylase and DOPA decarboxylase. The disease presents with three cardinal symptoms: akinesia (reduced mobility), rigidity (Muscle stiffness), and resting tremor. Dopamine does not cross the blood-brain barrier and cannot be administered directly as a medication. Management strategies for Parkinsonism include:

✵ replacement therapy using dopamine precursor drugs (DOPA derivatives) such as levodopa, madopar, nakom, etc.;

✵ inhibition of dopamine inactivation using monoamine oxidase inhibitors (deprenyl, nialamide, pyrazidol, etc.).

Depressive states are frequently associated with reduced levels of dopamine and norepinephrine in Nerve Cells.

Dopamine hypersecretion in the temporal lobe of the brain is observed in Schizophrenia.