Reductive amination NADH regeneration

In contrast, amino acid dehydrogenases comprise a well-known class of enzymes with industrial apphcations. An illustrative example is the Evonik (formerly Degussa) process for the synthesis of (S)-tert-leucine by reductive amination of trimethyl pyruvic acid (Scheme 6.12) [27]. The NADH cofactor is regenerated by coupling the reductive amination with FDH-catalyzed reduction of formate, which is added as the ammonium salt. 

An elegant four-enzyme cascade process was described by Nakajima et al. [28] for the deracemization of an a-amino acid (Scheme 6.13). It involved amine oxidase-catalyzed, (i )-selective oxidation of the amino acid to afford the ammonium salt of the a-keto acid and the unreacted (S)-enantiomer of the substrate. The keto acid then undergoes reductive amination, catalyzed by leucine dehydrogenase, to afford the (S)-amino acid. NADH cofactor regeneration is achieved with formate/FDH. The overall process affords the (S)-enantiomer in 95% yield and 99% e.e. from racemic starting material, formate and molecular oxygen, and the help of three enzymes in concert. A fourth enzyme, catalase, is added to decompose the hydrogen peroxide formed in the first step which otherwise would have a detrimental effect on the enzymes. 

Enzymatic synthesis of E-tm-leucine is another example of the use of isolated enzymes (Bommarius et al, 1995). An NADH-dependent leucine dehydrogenase was used as a catalyst for the reductive amination of the corresponding keto acid together with formate dehydrogenase (FDH) and formate as a cofactor regenerator (Fig. 19.5b Shaked and Whitesides, 1980 Wichmann et al, 1981). Furthermore, a unique membrane reactor system involving FDH and PEG-modihed-NAD for continuous NADH regeneration... 

Fig. 8 Synthesis of amino acids by a multienzyme system consisting of leucine dehydrogenase (LeuDH) catalyzing the reductive amination of the corresponding keto acid, L-lactate dehydrogenase (l-LDH), and lactate for the regeneration of NADH and urease for the in situ generation of ammonia. The coenzyme NAD+ was covalently bond to dextran, enzymes and dextran-coupled NAD+ were...

Because of its bulky, inflexible, and hydrophobic side chain, terf-leucine (2-amino-3,3-dimethylbutanoic acid, Tie) is an important amino acid used as template or catalyst compound in asymmetric synthesis and in peptidic medicinal compounds. L-Tle has attracted much attention as a key component of newly emerged drugs or as building block of ligands, catalysts, and auxiliaries for asymmetric synthesis. It is synthesized in ton-scale by reductive amination of trimethylpyruvic acid by means of LeuDH from Bacillus stearothermophilus with very high yield and excellent optical purity [153]. NADH, which is consumed during the reaction, can be regenerated by FDH from C. boidinii (Fig. 35). 

L-6-Hydroxynorleucine is a key intermediate used for the synthesis of a vasopepti-dase inhibitor. It was synthesized from 2-keto-6-hydroxyhexanoic acid by reductive amination using beef liver GluDH and GDH from Bacillus sp. for regeneration of NADH (Fig. 36) [155]. The educt of the reaction, 2-keto-6-hydroxyhexanoic acid is in equilibrium with 2-hydroxytetrahydropyran-2-carboxylic acid. 

Allysine ethylene acetal is synthesized from the corresponding keto acid by reductive amination using phenylalanine dehydrogenase (PDH) from Thermoacti-nomyces intermedius ATCC 33205 combined with FDH from C. boidinii SC13822 for the regeneration of NADH (Fig. 39). 

Figure 11. Hydrogenase-catalyzed NADH regeneration coupled to the reductive amination of a-ketoglutarate to L-glutmate catalyzed by L-glutamate dehydrogenase.

A very promising process route is the reductive amination of prochiral a-keto acids to a-amino acids with AADHs and the cofactor NADH and its regeneration by cooxidation of formate to CO2 by formate dehydrogenase (Fig. 15.3-1). 

FDH from C. boidinii was introduced by Whitesides and Shaked, and by Knla, Wandrey, and coworkers ° for regeneration of NADH. The advantages of this enzyme reaction are that the product CO2 is easy to remove, and the negative reduction potential ( = -0.42 v) for the FDH reaction drives the reductive amination to completion. 

Fig. 3. (A) Enzymatic synthesis of chiral synthon for Omapatrilat (1) Reductive amination of keto acid acetal (8) to (S)-allysine ethylene acetal (Z) by phenylalanine dehydrogenase. Regeneration of NADH was carried out using formate dehydrogenase. (B) Enzymatic synthesis of chiral synthon for Omapatrilat (1) Conversion of disulfide (U) to thiazepine (9) by S-lysine a-aminotransferase.

Figure 4 Reductive amination of 2-oxo-3,3-dimethyl butanoic acid with NADH regeneration (LeuDH, leucine dehydrogenase FDH, formate dehydrogenase).

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