Broad transaminase

Turning to l-AAO, Pantaleone s industrial research group have reported" on the properties and use of an l-AAO from Proteus myxofaciens, overexpressed in Escherichia coli This l-AAO, unusually, appears not to produce H2O2 in the catalytic reaction, thus making the addition of catalase unnecessary. The enzyme has a broad specificity, with a preference for nonpolar amino acids. This l-AAO was used in conjunction with a D-amino acid transaminase (d-AAT) and an alanine racemase (AR) to allow an efficient conversion of L-amino acid in to D-amino acid (Scheme 4). 

This enzyme [EC 2.6.1.1] (also known as transaminase A, glutamicioxaloacetic transaminase, and glutamic aspartic transaminase) catalyzes the reversible reaction of aspartate with a-ketoglutarate to produce oxaloace-tate and glutamate. Pyridoxal phosphate is a required cofactor. The enzyme has a relatively broad specificity, and tyrosine, phenylalanine, and tryptophan can all serve as substrates. 

Granulomatous hepatitis, with a rash or isolated increases in semm transaminase activities, can occur in patients with lung reactions (65). In the protracted acute and in chronic lung reactions hver injury (such as chronic active hepatitis) is more frequent than in acute reactions. Such cases usually show a broad spectrum of serological autoimmune reactions (lupus-like syndrome) (66). 

L-Amino acid transaminases are ubiquitous in nature and are involved, be it directly or indirectly, in the biosynthesis of most natural amino acids. All three common types of the enzyme, aspartate, aromatic, and branched chain transaminases require pyridoxal 5 -phosphate as cofactor, covalently bound to the enzyme through the formation of a Schiff base with the e-amino group of a lysine side chain. The reaction mechanism is well understood, with the enzyme shuttling between pyridoxal and pyridoxamine forms [39]. With broad substrate specificity and no requirement for external cofactor regeneration, transaminases have appropriate characteristics to function as commercial biocatalysts. The overall transformation is comprised of the transfer of an amino group from a donor, usually aspartic or glutamic acids, to an a-keto acid (Scheme 15). In most cases, the equilibrium constant is approximately 1. 

The use of liver function tests in the diagnosis and management of cirrhosis is discussed in the following sections. It may be useful to group the tests into two broad categories markers of hepatocyte damage such as the transaminases and markers of hepatocellular synthetic function, prothrombin time and albumin. 

The synthesis of chiral a-amino acids starting from a-keto acids by means of a transamination has been reported by NSC Technologies [26, 27]. In this process, which can be used for the preparation of l- as well as D-amino acids, an amino group is transferred from an inexpensive amino donor, e.g., L-glutamic acid, l-22, or L-aspartic acid, in the presence of a transaminase (= aminotransferase). This reaction requires a cofactor, most commonly pyridoxal phosphate, which is bound to the transaminase. The substrate specificity is broad, allowing the conversion of numerous keto acid substrates under formation of the L-amino acid products with high enantioselectivities [28]. 

Among the various enzymes capable of producing optically-active amino acids, transamination reactions, catalyzed by enzymes known as aminotransferases or transaminases, have broad potential for the synthesis of a wide variety of enantio-merically pure (R)- and (S)-compounds containing amine groups. Indeed, various examples of the use of aminotransferases for the production of d- and L-amino acids, both naturally-occurring and non-natural, have been published17 151. In addition, certain aminotransferases have been found to act on amines, and methods for the production of enantiomerically pure amines by transamination have been described116-211. This method allows for yields of up to 100% whereas routes based on hydrolases require external racemization to reach such yield levels. In this section we will focus on the application of aminotransferases. 

L-Phosphinothricin, the active ingredient of the broad-spectrum herbicide Basta (AgrEvo), can be obtained through enzymatic transamination of the corresponding oxoacid, 2-oxo-4-[(hydroxy)(methyl)phosphinoyl]butyric acid, in a coupled system with aspartate aminotransferase (AAT) and 4-aminobutyrate 2-ketoglutarate transaminase (E.C. 2.6.1.19) from E. coli (Fig. 12.7-6)[37 . In solutions containing 10% substrate, 85 % conversion was reached with only < 3 % amino acid by-products. For... 

The effectiveness of decarboxylation in driving the reaction to completion was demonstrated in a coupled enzymatic process by using phenylpyruvate as the starting 2-keto add. In this experiment, phenylpyruvate sodium salt and L-aspartate were incubated with E. coli broad-range transaminase at room temperature and pH... 

The pH-rate profile for the reaction catalyzed by the E. coli broad-range transaminase was determined using the immobilized transaminase with p-fluorophe-... 

Transaminase, broad-range, D-specific Origin microorganism, rec. in E. coli BioCatalytics AT-103... 

PLP is the coenzyme of more than 160 enzymes catalyzing a broad variety of reactions, predominantly involving an amino acid, an amine, or an oxoacid. Some of the selective examples are (1) the transfer of the alpha amine group from an amino acid to a ketoacid, catalyzed by transaminases, (2) the elimination or replacement of the substituent in the beta position of an amino acid or other compounds, catalyzed by lyases and synthases, respectively, and (3) the removal of the alpha carboxylic group from amino acids or other compounds, catalyzed by decarboxylases. More complex reactions are also carried out, involving, for example. 

The availabihty of a broad variety of oo-TAs together with efficient techniques to shift the equilibrium allows the biocatalytic synthesis of amines from the corresponding ketones via amino-group transfer. The potential of this protocol is demonstrated by a selection of amines, which can be obtained in nonracemic form by using the most prominent oo-transaminases (Scheme 2.227). 

Several metabolic blocks could account for the biochemical distortion observed in maple syrup disease. A deficiency in amino oxidase could lead to accumulation of amino acids. Because the enzyme has such a broad specificity, whenever it is completely deleted a more complex aminoaciduria can be expected to develop. The deletion of a specific transaminase could hardly explain the keto acid accumulation. Therefore, it seems more likely that the metabolic block involves a step between the keto acid and the simple acids, possibly the oxidative decarboxylation of the keto acid. This reaction requires coenzyme A, NAD, lipoic acid, and thiamine pyrophosphate, and it was described in some detail in the chapter devoted to the bioenergetic pathways. Leukocytes of at least some patients with maple syrup disease have been shown to contain normal transaminase activity but are defective in the oxidative decarboxylase. 

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