Alanine transaminase pyridoxal phosphate

This enzyme [EC 2.6.1.2], also known as glutamic-pyruvic transaminase and glutamic-alanine transaminase, catalyzes the pyridoxal-phosphate-dependent reaction of alanine with 2-ketoglutarate, resulting on the production of pyruvate and glutamate. 2-Aminobutanoate will also react, albeit slowly. There is another alanine aminotransferase [EC 2.6.1.12], better known as alanine-oxo-acid aminotransferase, which catalyzes the pyridoxal-phosphate-dependent reaction of alanine and a 2-keto acid to generate pyruvate and an amino acid. See also Alanine Glyoxylate Aminotransferase... 

This enzyme [EC 2.6.1.21], also known as D-aspartate aminotransferase, D-amino acid aminotransferase, and D-amino acid transaminase, catalyzes the reversible pyridoxal-phosphate-dependent reaction of D-alanine with a-ketoglutarate to yield pyruvate and D-glutamate. The enzyme will also utilize as substrates the D-stereoisomers of leucine, aspartate, glutamate, aminobutyrate, norva-hne, and asparagine. See o-Amino Acid Aminotransferase... 

In all cases the keto acids seem to be formed by typical a-ketogluta-rate-linked, pyridoxal phosphate-dependent transaminases (EC 2.6.1.6, etc.) (9, 154, 156, 157). There has been little study of isolated, presumably specific enzymes in connection with flavors, although the leucine and alanine aminotransferases of tomato have been precipitated with (NH4)oS04 (164, 165). Transaminase activity in Saccharomyces cere-visiae has a pH optimum of 7.2 (154), and a-ketoglutarate is the only amino group recipient (154, 166). Only aspartate and amino acids with hydrophobic side chains are acted on (154). 

D. Alanine aminotransferase (transaminase) requires pyridoxal phosphate. 

Determination of ALT ALT (formerly glutamate pyruvate transaminase) catalyzes the equilibrium transfer reaction of the amino group from L-alanine to 2-oxoglutarate to form L-glutamate and pyruvate ALT requires pyridoxal phosphate as coenzyme, which acts as an amino carrier. It is found in the main organs, such as the liver, kidney, and heart. The ALT activity in serum is elevated in diseases of the liver. 

Kynurenic acid and xanthurenic acid, side products of the reaction, are the products of the transamination of the a-amino group of kynurenine and 3-hydroxy-kynurenine to a-ketoglutaric acid in the presence of pyridoxal phosphate and an enzyme found in mammalian liver and kidney, kynurenine transaminase. The keto acid resulting from the transamination reaction condenses spontaneously. Liver homogenate also decarboxylates 3-hydroxykynurenine to yield 4,8-de-hydroxyquinoline. Kynurenase may catalyze the cleavage of the side chain of kynurenine or 8-hydroxy-kynurenine and lead to the formation of alanine and... 

Chloroalanine has been found to be an irreversible inhibitor of the pyridoxal phosphate-linked yS-aspartate decarboxylase/ aspartate aminotransferase/ and alanine racemase. The mechanism of inhibition is shown above by Eq. (7) (the sulfate reacts in the same manner) amino-ethane sulfonate irreversibly inhibits pyridoxal phosphate-linked GABA transaminase and L-serine-O-sulfate irreversibly inhibits aspartate aminotransferase. ... 

The function of a coenzyme is well illustrated by the role of pyridoxal phosphate in the transfer of amino groups. The enzyme alanine aminotransferase glutamate-pyruvate transaminase) catalyses the reaction of glutamate with pyruvate to form 2-oxoglutarate and alanine. In this reaction, the amino group of glutamate is transferred first to pyridoxal phosphate and then to pyruvate with the formation of alanine. 

The best method for the assay of pyridoxal phosphate is the use of tyrosine decarboxylase as described by Gunsalus, Bellamy, and Umbreit. The enzyme is prepared from a dried powder of cells of S. faecalis R. which has been grown deficient in vitamin Be by growth In a vitamin-Be-free alanine-rich medium. Thus, the decarboxylase is obtained almost completely resolved. This is a convenient preparation, since such a powder is stable for long periods and since the resolution of transaminases, decarboxylases, and tryptophanases isolated from tissues is a rather difficult task. The assay is performed manometrically by measuring the rate of CO2 liberation from tyrosine by the dried powder in the presence of pyridoxal phosphate. The rate of CO2 evolution is a function of the concentration of pyridoxal phosphate. 

Merewialis pereimu.—When aliphatic aldehydes are fed to this plant they are converted into the corresponding primary amines. A transaminase has been isolated which catalyses the transamination between a-amino-acids and aldehydes. L-Alanine was the most efficient amino donor, and all aldehydes in the homologous series from ethanal to undecanal were active amino acceptors. No requirement for pyridoxal phosphate could be demonstrated. In a later publication it was shown that this reaction takes place in many flowering plants. 

The recombinant (R)-transaminase prepared from cell extracts of rec E. coli SC16577 (40 g wet cells/200 ml suspension) was suspended in buffer containing 50 mM potassium phosphate pH 7,1 mM dithiothreitol, and 10 xM pyridoxal phosphate using a microfluidizer at 12000 psi followed by separation of cell debris. The reaction mixture contained 10 mL of this cell extract, keto acid 90 (6.0 g), D,L-alanine (12.0 g), pyridoxal phosphate (2.6 mg), dithiothreitol (15.0 mg) and 0.1 M potassium phosphate buffer pH 7.5 (80 mL). After incubating at 30°C with gentle shaking at 50 rpm for 39 h, the reaction was complete with 95% in-process yield and the product (R)-88a was isolated in 79% yield (5.4g) with >99% ee [155]. 

Various pyridoxal phosphate dependent enzymes compete with each other for the available pool of coenzyme. Thus the extent to which an enzyme is saturated with its coenzyme provides a means of assessing the adequacy of the body pool of coenzyme. This can be determined by measuring the activity of the enzyme before and after the activation of any apoenzyme present in the sample by incubation with pyridoxal phosphate added in vitro. Erythrocyte aspartate and alanine transaminases are both commonly used the results are usually expressed as an activation coefficient— the ratio of activity with added coenzyme to that without. 

Transaminases also belong to the group of transferases and catalyze desamination and amination reactions. Most amino acids are in equilibrium with the corresponding 0x0-carboxylic acid like alanine and pyruvate. The coenzyme of enzymatic transamination is pyridoxal-5 -phosphate 93, it is one of the most important coenzymes and is in effect the biologically active form of vitamin B6, which is converted to pyridoxamine 94 in this reaction. Besides transamination, pyridoxal-5 -phosphate 93 is also involved in decarboxylation and racemization processes. The aldehyde function of pyridoxal is reacting with the amine of an amino acid to form a Schiff base and thus invers-ing the acidity of the a-CH. The reversible transfer of an... 

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