Broad Specificity Amino Acid Racemase

The one-base mechanism is characterized by the retention of the substrate-derived proton in the product (internal retum).30) With this criterion, reactions catalyzed by a-amino-c-caprolactam racemase,323 amino acid racemase of broad specificity from Pseudomonas striata333 have been considered to proceed through the one-base mechanism. However, such internal returns were not observed in the reactions of alanine racemases from K coli B,33) B. stearothermophilus,263 and S. typhirmaium (DadB and /1/r).263 The internal return should not be observed in the two-base mechanism, because the base catalyzing the protonation to the intermediate probably obtains the proton from the solvent. But the failure of the observation of the internal return can be also explained by the single-base mechanism in which exchange of the proton abstracted from the substrate a-carbon with the solvent is much faster than its transfer to the a-carbanion. Therefore, lack of the internal return does not directly indicate the two-base mechanism of the alanine racemase reaction. 

An amino acid racemase which shows very broad substrate specificity was discovered in Pseudomonas striata (= Ps. putida), purified, and characterized1 91. The enzyme catalyzes racemization of various amino acids except aromatic and acidic... 

Racemic a-amino amides and a-hydroxy amides have been hydrolyzed enantio-selectively by amidases. Both L-selective and o-selective amidases are known. For example, a purified L-selective amidase from Ochrobactrum anthropi combines a very broad substrate specificity with a high enantioselectivity on a-hydrogen and a,a-disubstituted a-amino acid amides, a-hydroxyacid amides, and a-N-hydroxya-mino acid amides [102]. A racemase (a-amino-e-caprolactam racemase, EC 5.1.1.15) converts the o-aminopeptidase-catalyzed hydrolysis of a-amino acid amides into a DKR (Figure 6.38) [103]. 

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). 

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