Hydroxylation of the aromatic amino acids

Scheme 1. Hydroxylation of the aromatic amino acids ( -Phe, L-Tyr, and L-Trp) as catalyzed by their respective hydroxylase enzymes (PAH, TH, TPH1, and TPH2) using the cofactor BH4 and dioxygen as additional substrates.

The structural similarity of the catalytic domains of the enzymes of the AAH family, together with the identical reaction that they catalyze (i.e., hydroxylation of aromatic substrates) and the common dependency on BH4 and 02 (Section I), suggests that the mechanisms by which these enzymes operate are similar. It is assumed that the general AAH reaction mechanism follows a two-step reaction route in which a high-valent iron-oxo (FeIV=0) complex is formed in the first step, and that this intermediate is responsible for the hydroxylation of the aromatic amino acid substrate in the second step (15,26-28,50). The first step starts with 02 binding and activation and proceeds via a Fe-0-0-BH4 bridge and a subsequent heterolytic cleavage of the... 

B. Step II—Feiv=0-Catalyzed Hydroxylation of the Aromatic Amino Acid... 

The important metaboUsm of the neurotransmitters norepinephrine, epinephrine, dopa, and serotonin involves pterin-dependent monooxygenases. The direct biocatalytic hydroxylation of the aromatic amino acids phenylalanine, tyrosine, and tryptophane requires tetrahydrobiopterin and Fe as the cofactors [60]. The cleavage of unsaturated glyceryl ethers by glyceryl ether monooxygenase also requires tetrahydrobioterin as the cofactor [61]. 

Scheme 5. The electronic rearrangements of the aromatic amino acid substrate hydroxylation.

Although regulation is unique for each member of the aromatic amino acid hydroxylase family, the catalytic mechanism and cofactor requirements for members of the family are identical. During the reactions of all three enzymes, the dioxygen molecule is cleaved and incorporated as a hydroxyl group into both the aromatic amino acid and BH4. Each enzyme in the family displays its own unique substrate specificity profile. Two interesting questions about this enzyme family relate to the actual hydroxylation mechanism and how enzyme activity is altered by changes in BH4 levels. Problems in any one of these hydroxylation systems can arise from either an inadequate supply of the BH4 cofactor or a defect in the enzyme or its expression. 

The quinone ring is derived from isochorismic acid, formed by isomerization of chorismic acid, an intermediate in the shikirnic acid pathway for synthesis of the aromatic amino acids. The first intermediate unique to menaquinone formation is o-succinyl benzoate, which is formed by a thiamin pyrophosphate-dependent condensation between 2-oxoglutarate and chorismic acid. The reaction catalyzed by o-succinylbenzoate synthetase is a complex one, involving initially the formation of the succinic semialdehyde-thiamin diphosphate complex by decarboxylation of 2-oxoglutarate, then addition of the succinyl moiety to isochorismate, followed by removal of the pyruvoyl side chain and the hydroxyl group of isochorismate. 

Most aromatic hydroxylases aue either cytochrome- or flavin-dependent enzymes the three enzymes that catadyze hydroxylation of the anomatic amino acids phenylalanine, tyrosine, amd tryptophaui ane apparently unique in... 

The pterin-dependent oxygenases, typified by the aryl amino acid hydroxylases, are a small family of closely related enzymes, which are essential to mammalian physiology. This class of metalloenzymes employs tetrahydrobiopterin (BH4) as a two-electron donating cofactor for the activation of O2. Members of this class include phenylalanine (PheH), tyrosine (TyrH) and tryptophan (TrpH) hydroxylases, which effect regiospecific aromatic hydroxylations of the namesake amino acids. 

The amino acids with nonpolar, aliphatic side chains, Ala, lie. Leu, Met, and Val, are sufficiently hydro-phobic that they are most often buried in the generally hydrophobic core of non-membrane-embedded proteins. Note that lie and Val have particularly sterically hindered P-carbons. Of the aromatic amino acids. His, with a pK of around 6, will mostly be in the uncharged form at physiological pH values (therefore more often hydrophobic than polar), and will be a likely choice for reactions which involve proton transfer. Phe and Trp are clearly hydrophobic. Despite having a polar hydroxyl group, if we consider the free energy required to transfer... 

Subsequent studies by the same group (W7) have shown that the increased endogenous fluorescence in patients with chronic renal failure is due to the unconjugated pteridine, xanthopterin (2-amino-4,6-pteridinedione, 179 Da see Table 4 and Figs. 10 and 11) (B23, Dll, L15, Pll). Unlike the conjugated pteridines (folates), the function of many of the unconjugated pteridines (pterins) has yet to be elucidated (U2, Z3). So far only biopterin has been shown to have a defined role, being a cofactor in the hydroxylation of several aromatic amino acids involved in the formation of neuronal hormones such as catecholamines and serotonin. 

Since the phenylethylamines 312 produced by these decarboxylases are substrates for systems containing dopamine jS-hydroxylase (EC 1.14.17.1), the availability of 7>R and 3S isotopically labeled samples of the aromatic amino acids has allowed the stereochemistry of the hydroxylation of dopamine 313 to yield norepinephrine 314 to be studied (Scheme 83). It was shown that the 3-pro-S hydrogen, Hg, was lost from phenylalanine 297a in the process and that the hydroxylation yielding 314 therefore occurred with retention of configuration (319). 

Dalgliesh, C.E. Nonspecific Formation of Hydroxylated Metabolites of the Aromatic Amino Acids ... 

Tyrosine or phenylalanine residues in position 2 are essential for activity. Replacement of the aromatic amino acid by serine inactivates the hormone completely. The phenylalanine derivative is less active than the tyrosine derivative, indicating that the hydroxyl group of tyrosine, although not essential, enhances the hormonal activity. 

Origin of the side chain of the aromatic amino acids and of prephenic acid from a three-carbon glycolytic intermediate was established by isotopic results as previously mentioned. Evidence of this is that the / -carbon of tyrorine is derived about equally and almost entirely from C-1 and 6 of glucose (217), and that the o-carbon was h hly labeled while the /3-carbon was unlabeled in tyrorane derived from pyruvate-a-C (219). Synthesis of prephenic acid presumably results from condensation of a pyruvate moiety with a cyclic intermediate beyond shikimic acid that has lost two hydroxyl groups. 

In the sirodesmins (see Table III), the structure of the aromatic amino acid precursor has been obscured by prenylation, hydroxylation, and ring... 

The conversion of the aromatic amino acid phenylalanine to tyrosine is one of the most well-studied aerobic hydroxylation reactions. The reaction, itself, serves a dual role in mammalian metabolism. It is, first of all, an obligatory step in the combuston of phenylalanine to COg and water, there being no alternate pathway in animals for the cleavage of the benzene ring. Secondly, the reaction provides an endogenous source for the amino acid tyrosine. It is because this reaction occurs that tyrosine is classified as a nonessential amino acid. 

The liver is also the principal metabolic center for hydrophobic amino acids, and hence changes in plasma concentrations or metabolism of these molecules is a good measure of the functional capacity of the liver. Two of the commonly used aromatic amino acids are phenylalanine and tyrosine, which are primarily metabolized by cytosolic enzymes in the liver [1,114-117]. Hydroxylation of phenylalanine to tyrosine by phenylalanine hydroxylase is very efficient by the liver first pass effect. In normal functioning liver, conversion of tyrosine to 4-hy-droxyphenylpyruvate by tyrosine transaminase and subsequent biotransformation to homogentisic acidby 4-hydroxyphenylpyruvic acid dioxygenase liberates CO2 from the C-1 position of the parent amino acid (Fig. 5) [1,118]. Thus, the C-1 position of phenylalanine or tyrosine is typically labeled with and the expired C02 is proportional to the metabolic activity of liver cytosolic enzymes, which corresponds to functional hepatic reserve. Oral or intravenous administration of the amino acids is possible [115]. This method is amenable to the continuous hepatic function measurement approach by monitoring changes in the spectral properties of tyrosine pre- and post-administration of the marker. 

Some rather important indole derivatives influence our everyday lives. One of the most common ones is tryptophan, an indole-containing amino acid found in proteins (see Section 13.1). Only three of the protein amino acids are aromatic, the other two, phenylalanine and tyrosine being simple benzene systems (see Section 13.1). None of these aromatic amino acids is synthesized by animals and they must be obtained in the diet. Despite this, tryptophan is surprisingly central to animal metabolism. It is modified in the body by decarboxylation (see Box 15.3) and then hydroxylation to 5-hydroxytryptamine (5-HT, serotonin), which acts as a neurotransmitter in the central nervous system. 

Dopamine is the decarboxylation product of DOPA, dihydroxyphenylalanine, and is formed in a reaction catalysed by DOPA decarboxylase. This enzyme is sometimes referred to as aromatic amino acid decarboxylase, since it is relatively non-specific in its action and can catalyse decarboxylation of other aromatic amino acids, e.g. tryptophan and histidine. DOPA is itself derived by aromatic hydroxylation of tyrosine, using tetrahydrobiopterin (a pteridine derivative see Section 11.9.2) as cofactor. 

The aromatic amino acids, phenylalanine, tyrosine, and tryptophan, have ring structures and are nonpolar with the exception of the hydroxyl group of tyrosine. 

Among the essential amino acids, the aromatic amino acids (phenylalanine, tyrosine, and tryptophan) form by a pathway in which chorismate occupies a key branch point. Phosphoribosyl pyrophosphate is a precursor of tryptophan and histidine. The pathway to histidine is interconnected with the purine synthetic pathway Tyrosine can also be formed by hydroxylation of phenylalanine (and thus is considered conditionally essential). The pathways for the other essential amino acids are complex. 

Reaction Lx x ill. Direct Replacement of the Aromatic Amino-group by Hydroxyl. (B., 7, 77, 809 D.R.P., 109102.)—The simple primary amino-groups in the benzene series are not easily replaced directly by hydroxyl unless an activating group (e.g., N02) be present in the o- or p-position. a-Naphthols, however, are readily obtained by heating a-naphthylamine derivatives with fairly concentrated acid under pressure. 

BA biosynthesis begins with the conversion of tyrosine to both dopamine and 4-hydroxyphenylacetaldehyde by a lattice of decarboxylations, orfho-hydroxylations, and deaminations.1 The aromatic amino acid decarboxylase (TYDC) that converts tyrosine and dopa to their corresponding amines has been purified, and several... 

The fourth class, the pterin-dependent hydroxylases, includes the aromatic amino acid hydroxylases, which use tetrahydrobiopterin as cofactor for the hydroxylation of Phe, Tyr, and Trp. The latter two hydroxylases catalyse the rate-limiting steps in the biosynthesis of the neurotransmitters/hormones dopamine/noradreanalme/ adrenaline and serotonin, respectively. 

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