Amino acid amidase

FIGURE 19.1 General reactions of hydrolase enzymes. In the same group as nitrilase enzymes are the amidases. This includes amino acid amidase ... 

DSM developed a slightly different approach towards enantiopure amino acids. Instead of performing the Strecker synthesis with a complete hydrolysis of the nitrile to the acid it is stopped at the amide stage. Then a stereoselective amino acid amidase from Pseudomonas putida is employed for the enantioselective second hydrolysis step [83], yielding enantiopure amino acids [34, 77, 78]. Although the reaction is a kinetic resolution and thus the yields are never higher than 50% this approach is overall more efficient. No acylation step is necessary and the atom efficiency is thus much higher. A drawback is that the racemisation has to be performed via the Schiff s base of the D-amide (Scheme 6.23). 

Recently it was reported that an a-amino-e-caprolactam racemase from Achro-mobacter obae can racemise a-amino acid amides efficiently. In combination with a D-amino acid amidase from Ochrobactrum anthropi L-alanine amide could be converted into D-alanine. This tour de force demonstrates the power of the racemase [84]. If racemic amide is used as a starting material the application of this racemase in combination with a d- or L-amidase allows the preparation of 100% d- or L-amino acid, a dynamic kinetic resolution instead of DSM s kinetic resolution (Scheme 6.24). 

In the same group as nitrilase enzymes are the amidases. This includes amino acid amidase (EC 3.5.1.4), used to prepare amino acids, usually through resolution, and also penicillin G acylase (penicillin G amidohydrolase) (EC 3.5.1.11), used in the manufacture of semisynthetic penicillins [102,103]. Immobilized penicillin G acylase has most recently been used to catalyze the formation of A-a-phenylacetyl amino acids, which can then be used in peptide coupling reactions (see Section 13.2.3.2) [104]. 

Asano et al. have also purified a D-stereospecific amino acid amidase from another Ochrobadrum anthropi isolate137, 38i. Recently, a new amidase from Comamonas acidovorans has been reported that exhibits a broad substrate specificity and also d-amino acid amidase activity1391. In addition, a D-specific amidase has been identified in Arthrobacter sp. NJ-261401. In contrast to the D-selective enzymes of Ochrobadrum sp. and Comomonas acidovorans, the D-amide hydrolase identified in Arthrobacter sp. NJ-26 was very substrate specific a good hydrolysis rate was only observed for d-alanine amide. 

Recently the enzyme has been purified and thoroughly characterized. It was identified as an amino acid amidase, most probably belonging to the group of... 

Synthesis of (S)- and (/ )-a-amino acids using nitrile hydratase (NHase) from Rhodococcus opacus, a-amino-e-caprolactam racemase (ACL) from Achromobacter obae, and different amidases (amidase 1, o-amino-peptidase from Ochrobactrum anthropi amidase 2, i-amino acid amidase from Brevundimonas diminuta) [111, 112]. / =CH3, CH CH, CH3CH(CH3), CH3CH(CH3)CH2. 

Discovery of D-Aminopeptidase, D-Amino Acid Amidase, and Alkaline o-Peptidase... 

Since o-stereospecific amino acid amides and peptide hydrolases were previously unknown and were not targets of enz3unology, we started to screen for these enzymes and subsequently discovered three kinds that exhibited D-stereoselectivities for o-amino acid derivatives o-aminopeptidase [4], o-amino acid amidase [5], and alkaline o-peptidase [6] (Table 19.1). Ochrobactrum anthropi Cl-38 was isolated through an enridiment culture technique as a utilizer of o-alanine amide (o-Ala-NH ) as the sole... 

D-Amino acid amidase (DaaA) 0. anthropi SV3 Ser Dynamic kinetic resolution... 

Ochrobactrum anthropi SV3 was isolated as a D-valine amide degrader after a 4-month acclimation of bacterial culture. The strain produced an enzyme that was characterized as o-amino acid amidase (DaaA) [5], Furthermore, Bacillus cereus DF4-B excreted alkaline o-peptidase (ADP), which acted on the synthehc oligopeptide D-phenylalanine tetramer (o-Phe). The enzyme was characterized as the first endopeptidase acting on D-amino acid-containing peptides by recognizing the second amino acid from its N-terminus under alkaline conditions [6]. 

D-Amino Acid Amides by i-Amino Acid Amidase in the Presence of ACL Racemase... 

Komeda, H. and Asano, Y, "Gene cloning, nucleotide sequencing, and purification and characterization of the D-stereospecific amino-acid amidase from Ochrobactrum anthropi SV3." Eur.. Biochem., 267, 2028-2036 (2000). 

Okazaki, S., Suzuki, A., Komeda, H., Yamaguchi, S., Asano, Y, and Yamane, T, "Crystal structure and functional characterization of a u-stereospecific amino acid amidase from Ochrobactrum anthropi SV3, a new member of the penicillin-recognizing proteins." /. Mol. Biol, 368, 79-91 (2007). 

Asano, Y., Mori, T., Hanamoto, S., Kato, Y, and Nakazawa, A., "A new D-stereospecific amino acid amidase from Ochrobactrum anthropi." Biochem. Biophys. Res. Commun., 162, 470-474 (1989). 

Yamaguchi, S., Komeda, H., and Asano, Y., "New enzymatic method of chiral amino add synthesis by dynamic kinetic resolution of amino acid amides use of stereoselective amino acid amidases in the presence of a-amino-e-caprolactam racemase." Appl. Environ. Microbiol, 73,5370-5373 (2007). 

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