Phenylpyruvic acid oxidase

Tyrosyluria 4-Hydroxyphenylpyruvic, 4-hydroxyphenyl-lactic and 4-hydroxyphenylacetic acids Tyrosine Delayed maturation of 4-hydroxy-phenylpyruvic acid oxidase (EC 1.13.11.27) 16.5.2... 

R. Fernandez-Lafuente, V. Rodriguez, and J.M. Guisan, The coimmobilization of D-amino acid oxidase and catalase enables the quantitative transformation of D-amino acids (D-phenylalanine) into alpha-keto acids (phenylpyruvic acid). Enz. Microb. Technol. 23, 28-33 (1998). 

L-Amino acid oxidase has been used to measure L-phenylalanine and involves the addition of a sodium arsenate-borate buffer, which promotes the conversion of the oxidation product, phenylpyruvic acid, to its enol form, which then forms a borate complex having an absorption maximum at 308 nm. Tyrosine and tryptophan react similarly but their enol-borate complexes have different absorption maxima at 330 and 350 nm respectively. Thus by taking absorbance readings at these wavelengths the specificity of the assay is improved. The assay for L-alanine may also be made almost completely specific by converting the L-pyruvate formed in the oxidation reaction to L-lactate by the addition of lactate dehydrogenase (EC 1.1.1.27) and monitoring the oxidation of NADH at 340 nm. 

Correct answer = B. Alkaptonuria is a rare metabolic disease involving a deficiency in homogentisic acid oxidase, and the subsequent accumulation of homogentisic acid in the urine, which turns dark upon standing. The elevation of methylmalonate (due to methylmalonyl CoA mutase deficiency), phenylpyruvate (due to phenylalanine hydroxlyase deficiency), a-ketoisovalerate (due to branched-chain a-ketoacid dehydrogenase deficiency), and homocystine (due to cystathionine synthase deficiency) are inconsistent with a healthy child with darkening of the urine. 

Additional errors of phenylalanine and tyrosine metabolism include tyrosinosis, or hereditary tyrosinemia, neonatal tyrosinemia, and alcaptonuria. In the first case, there is a probable defect in p-hydroxyphenylpyruvate oxidase. In neonatal tyrosinemia, the problem is transient and may be solved by the administration of ascorbic acid. Ascorbic acid is apparently a cofactor for p-hydroxy-phenylpyruvate oxidase. Alcaptonuria is a benign disorder in which homogen-tisic acid oxidase is inoperative and homogentisic acid is excreted in the urine. Air oxidizes the homogentisic acid to a pigment, giving urine a black color. This pigment also accumulates in the patient s tissues. 

Figure 8. Migration of the side chain during hydroxylation of phenylpyruvates with (hydroxy) phenyl-pyruvic acid oxidase...

Phenylalanine, synthesized from [ C]cyanide by the Bucherer modification of the Strecker synthesis, was resolved into its l- and D-iso-mers by the action of the d- and L-amino acid oxidases, respectively. The optically active amino acid was separated from phenylpyruvic acid by cation exchange chromatography (3). Similarly, DL-[ F]acyl-p-fluoro-phenylalanine has been subjected to stereospecific deacylation with the fungal enz)nne, L-amino acylase enz)nnatically generated L-[ F]p-fluoro-phenylalanine was separated from the D-acyl amino acid by column chromatography (4). 

Cinnamic acid (48) is synthesized universally in higher plants and widely in fungi from (25)-phenylalanine by phenylalanine ammonium lyase. This process is a trans elimination of the elements of ammonia, with stereospecific loss of the 3-pro S) proton of phenylalanine (46). Phenylpyruvic acid (49) (shown in its enol form) is the normal biogenetic precursor of phenylalanine, and is in equilibrium with it by means of the action of aminotransferases and amino acid oxidases (see Fig. 11). To distinguish between the participation of cinnamic acid (48) and phenylpyruvic acid (49) and to clarify the mechanism involved in the proton losses, (2R,35)-[3- H]-, (2S,3R)-[3- H], and (25)-[f/- C]phenylalanines were administered to the cultures. The incorporations resulted in the removal of 57% and 76%, respectively, of the labeled hydrogen. 

Experiments with (2S,3R)- and (2/, 35 )-[3- H]phenylalanines gave tritium retentions of 44% and 24%, respectively (Vederas and Tamm, 1976). Simultaneous incorporation of equal amounts of both enantiomers led to the expected 34% retention of hydrogen label. Transamination occurs stereo-specifically at position 2 of the amino acid therefore, the participation of at least two enzymes with different stereochemical requirements at the 3 position is reasonable. Two biosynthetic pathways are consistent with the data available (see Fig. 13). Path A in Fig. 13 depicts (2 S )-phenylalanine as the actual precursor which is in rapid equilibrium with its enantiomer in path B phenylpyruvic acid (49), derived directly from shikimic acid, is the primary precursor. Considerable suppression of the incorporation of D-amino acid by phenylpyruvic acid (49) indicated that the naturally abundant L-enantiomer is the actual primary precursor, thus demonstrating that path A (Fig. 13) is probably the main biosynthetic route. Both enantiomers are in rapid equilibrium with phenylpyruvic acid (49) via the action of aminotransferases or amino acid oxidases. The stereochemistry of hydrogen loss... 

Tyrosine Oxidation (p-Hydroxyphenylpyruvate Oxidase) (Eq. 17) The interruption of tyrosine degradation at the stage of p-hydroxy-phenylpyruvate (pHPP) in scurvy has long been considered Ae clearest evidence of a biochemical role for ascorbic acid. The intmmediate accumulates and is excreted when doses of tyrosine are given to scorbutic man, other primates, and guinea pigs, and to premature infants (K8). 

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