Hydrogenation amino acid substrates

The three-dimensional structure of the complex of D-amino acid oxidase with the substrate analog benzoate has been determined. The carboxyl group of the inhibitor is bound by an arginine side chain (Fig. 15-11) that probably also holds the amino acid substrate. There is no basic group nearby in the enzyme that could serve to remove the a-H atom in Eq. 15-26 but the position is appropriate for a direct transfer of the hydrogen to the flavin as a hydride ion as in Eq. 15-23.161/162/257 In spite of all arguments to the contrary the hydride ion mechanism could be correct However, an adduct mechanism is still possible. 

Figure 10. The mechanism of catalytic asymmetric hydrogenation of the amino acid substrates. This diagram depicts the structures of the intermediates. The absolute configurations refer to S,S-chiraphos.

The most versatile of the coenzymes is perhaps pyridoxal phosphate (PEP). The PEP containing enzymes catalyze a wide variety of reactions such as racemization, transamination, [3- and a-decarboxylation, and interconversion of side chains. The first step of all these reactions is the transition between an internal aldimine intermediate to an external aldimine intermediate, which involves the condensation of PEP with an external amino acid substrate to form a Schiff base. The internal aldimine intermediate can then either undergo a-decarboxylation to convert the amino acid substrate into amines and aldehydes, or lose the a-hydrogen... 

In each of these transformations, one of the bonds to the cr-carbon of the amino acid substrate is broken in the first step of the reaction. Decarboxylation breaks the bond joining the carboxyl group to the cr-carbon transamination, racemization, and o , 8-elimination break the bond joining the hydrogen to the cr-carbon and —Cp bond cleavage breaks the bond joining the R group to the a-carbon. 

After an initial a-hydrogen abstraction, transfonnalions at P-carbon or y-carbon of a-amino acids may occur. When a-amino acid substrates possess a substituent at either P- or y-position, they can function as good leaving groups such as COO , OH(CH3) or SH. Either elimination or replacement of the substituent yields corresponding oxo acids. In the case of reactions, labilization of P-hydrogen to form a P-carbanion intermediate is necessary. 

The choice of chloromethylated polystyrene as the resin support to bind the amino acid substrate by benzyl ester is amply justified on the basis of the frequent use of benzyl esters of amino acids in solution and methods of peptide synthesis (Greenstein and Winitz, 1961 Schroder and Lubke, 1965 Bodanszky and Ondetti, 1966). Such a bond is stable during the various reactions of peptide synthesis but is readily cleaved by anhydrous hydrogen bromide at the end of synthesis. Details for the improved synthesis of chloromethylated co(polystyrene-DVB) have been reported (Feinberg and Merrifield, 1974 see also Chapter 2). Both microporous (swellable) and macroreticular or macroporous polymers have been examined as support material (Tilak and Hollinden, 1971 Sano et aL, 1971), but in most synthetic reactions the swellable resins have an edge over the nonswellable resins. 

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]. 

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