Amino homophenylalanine

Hsu et have cloned two enzymes from Deimcoccus radiodurans for overexpression in E. coli in order to carry out a dynamic kinetic resolution to obtain L-homophenylalanine, frequently required for pharmaceutical synthesis. The starting material is the racemic mixture of A acetylated homophenylalanine, and the two enzymes are an amino acid A -acylase, which specifically removes the acetyl group from the L-enantiomer, and a racemase, which interconverts the D- and L-forms of the A acyl amino acids. The resolution was carried out successfully using whole-cell biocatalysts, with the two enzymes either expressed in separate E. coli strains or coexpressed in the same cells. 

A/-Carbamoylase Combined with Af-Acyl Amino Acid Racemase to Produce L-Homophenylalanine... 

In Section 5.03.6.2, a stereoselective synthesis of L-homophenylalanine from the racemic AAacetylated amino acid is described. The authors, however, found that substrate solubility limited the utility of this procedure. Having found an L-N-carbamoylase in Bacillus kaustophilus, they introduced the gene for this enzyme together with that for the N-acyl amino acid racemase from D. radiodurans into E. coli for coexpression. These cells, permeabilized with 0.5% toluene, were able to deliver L-homophenylalanine in 99% yield and were able to be used for multiple reaction cycles. 

Scheme 10.8 outlines the application of rhodium-catalyzed allyhc amination to the preparation of (il)-homophenylalanine (J )-38, a component of numerous biologically active agents [36]. The enantiospecific rhodium-catalyzed allylic amination of (l )-35 with the lithium anion of N-benzyl-2-nitrobenzenesulfonamide furmshed aUylamine (R)-36 in 87% yield (2° 1° = 55 1 >99% cee) [37]. The N-2-nitrobenzenesulfonamide was employed to facilitate its removal under mild reaction conditions. Hence, oxidative cleavage of the alkene (R)-36 followed by deprotection furnished the amino ester R)-37 [37, 38]. Hydrogenation of the hydrochloride salt of (l )-37 followed by acid-catalyzed hydrolysis of the ester afforded (i )-homophenylalanine (R)-3S in 97% overall yield. 

Although this enzymatic process fills only a niche in the L-lysine market, it is a successful example of a general method for amino acid resolution. It has some superior features compared to the Tanabe L-aminoacylase approach. The L-lysine can be extended to non-protein amino acids such as the use of P. putida aminopeptidase to resolve DL-homophenylalanine to produce precursors for the anti-hypertensive dmg Enalapril. A similar approach has also been used for the production of L-cysteine from DL-2-amino-A2-thiazohne-4-caiboxylate using Sarcina lucea, which is remarkable in that both isomers form L-cysteine. 

In the mid to late 1980s, many research groups focused on methods and processes to prepare L-phenylalanine (Chapter 3). This was a direct result of the demand for the synthetic, artificial sweetener aspartame. One of the many routes studied was the use of phenylalanine dH (Scheme 19.4, R = C6H5CH2) with phenylpyruvate (PPA) as substrate.57-58 This enzyme from Bacillus sphaericus shows a broad substrate specificity and, thus, has been used to prepare a number of derivatives of L-phenylalanine.59 A phenylalanine dH isolated from a Rhodococcus strain M4 has been used to make L-homophenylalanine (.S )-2-amino-4-pheny I butanoic acid], a key, chiral component in many angiotensin-converting enzyme (ACE) inhibitors.40 More recently, that same phenylalanine dH has been used to synthesize a number of other unnatural amino acids (UAAs) that do not contain an aromatic sidechain.43... 

In the active site of the enzyme PLP forms an internal aldimine (Schiff base) with Lys-270 (Fig. 9.13,1). When the substrate is bound at the active site, its a-amino group attacks the C-4 atom of the coenzyme and replaces the -amino group of Lys-270 from its bond with PLP. This transaldimination reaction probably proceeds via a tetrahedral intermediate (a gem-diamine). Spectral evidence for formation of a gem-diamine in this step has recently been obtained in studies of the reaction of tryptophanase with L-homophenylalanine.41 The gem-diamine is subsequently converted to an external, PLP-substrate aldimine, and the -amino group of Lys-270 is released (Fig. 9.13, II). The equilibrium constant of this step with L-tryptophan is determined to be 11.6 mM.78 ... 

Subsequently, shallow water collections of Lyngbya majuscula from Puerto Rico and the Dry Tortugas yielded additional supplies of ATX as well as a new congener termed antillatoxin B (Figure 6.10) [149]. The structure of the new metabolite was determined largely by comparison with the spectroscopic data set for ATX, and stereochemistry deduced by Marfey s analysis for L-alanine while the i-N-methyl homophenylalanine was proposed based on nuclear Overhauser effect (nOe) and bioassay results. Substitution of i-N-methyl homo-phenylalanine, an intriguing amino acid of quite rare occurrence in natural products, for i-N-methyl valine... 

The synthesis of the angiotensin-converting enzyme (ACE) inhibitor enalapril (1) incorporates the natural amino acids L-alanine and L-proline [1,2]. Although the compound can be thought of as a derivative of homophenylalanine, this part of the compound is prepared by a reductive amination with the keto ester (2), while the amino acids are coupled through an A-carboxy anhydride 3 (Scheme 1) [3-7]. 

A variety of commercially important Angiotensin Converting Enzyme (ACE) inhibitors contain an (S)-homophenylalanine moiety which can be introduced starting from various building blocks [1], One of the most useful is ethyl (R)-2-hy-droxy-4-phenylbutyrate (HPB ester) 1 which after introduction of a leaving group can be coupled with the respective amino moiety with inversion of the stereogenic center (Fig. 1). Since the patents for several ACE inhibitors have already expired or will soon do so, the production costs will become very important. This calls for the development of more efficient syntheses both for the ACE inhibitors as well as for key intermediates. In this chapter we will describe four technically feasible routes for the synthesis of 1 that have been developed by us in the last few years... 

A typical example for an efficient transamination process is the production of l-alanine, l-25, which is carried out in a continuous manner starting from pyruvate, 24, and L-glutamate, l-22, with a high space-time yield of 4.8kg/(L-d) (Fig. 13) [28], In addition, several non-proteinogenic a-amino acids, e.g., L-phosphinothri-cine, L-homophenylalanine, and L-tert-leucine have been also produced via transamination. 

Enantiomerically pure a-H-amino acids are intermediates in the synthesis of antibiotics used for parenteral nutrition and for food and feed additives (see also Chapter 12.2). Examples are D-phenylglycine and 4-hydroxyphenylalanine for semisynthetic (3-lactam antibiotics and L-phenylalanine for the peptidic sweetener aspartame. DSM used this process to produce also L-homophenylalanine, a potential precursor molecule for several ACE-inhibitors. 

The N-acetyl-D,L-amino acid precursors are conveniently accessible through either acetylation of D,L-amino acids with acetyl chloride or acetic anhydride in a Schotten-Baumann reaction or via amidocarbonylation I801. For the acylase reaction, Co2+ as metal effector is added to yield an increased operational stability of the enzyme. The unconverted acetyl-D-methionine is racemized by acetic anhydride in alkali, and the racemic acetyl-D,L-methionine is reused. The racemization can also be carried out in a molten bath or by an acetyl amino acid racemase. Product recovery of L-methionine is achieved by crystallization, because L-methionine is much less soluble than the acetyl substrate. The production is carried out in a continuously operated stirred tank reactor. A polyamide ultrafiltration membrane with a cutoff of 10 kDa retains the enzyme, thus decoupling the residence times of catalyst and reactants. L-methionine is produced with an ee > 99.5 % and a yield of 80% with a capacity of > 3001 a-1. At Degussa, several proteinogenic and non-proteinogenic amino acids are produced in the same way e.g. L-alanine, L-phenylalanine, a-amino butyric acid, L-valine, l-norvaline and L-homophenylalanine. 

Recently (12), the acid form of ultrastable zeolite Y (H-LFSY) has been studied as a solid catalyst in the reaction of a-amino acids with methanol at 100-130 °C (15-20 bar). The yields of the amino acids were DL-homophenylalanine, 68%, D-phenylglycine, 86%, L-phenylalanine, 77%, and D-p-hydroxy-phenylglycine, 14%. 

Figure 2 shows the effect of refluxing time on the conversion of amino acid homophenylalanine. At first, increasing reaction times increases the reaction conversion as expected. However, extending the reaction time beyond 5 hours decreases the conversion. Possible side reactions may occur at this moment. 

In the oxidative deamination reaction, the enzyme was active toward N-[l-D-(carboxyl)ethyl]-L-methionine, N-[l-D-(carboxyl)ethyl]-L-phenylalanine, etc. The substrate specificity for amino donors of ODH in the reductive secondary amine-forming reaction was examined with pyruvate as a fixed amino acceptor [15,24]. The enzyme utilized L-norvaline, L-2-aminobutyric acid, L-norleucine, P-chloro-L-alanine, o-acetyl-L-serine, L-methionine, L-isoleucine, L-valine, L-phenylalanine, L-homophenylalanine, L-leucine, L-alanine, etc. 3-Aminobutyric acid and L-phenylalaninol also acted as substrates for the enzyme. Other amino compounds, such as P-amino acids, amino acid esters and amides, amino alcohols, organic amines, hydroxylamines, and hydrazines, were inactive as substrates. Pyruvate, oxaloacetate, glyoxylate, and a-ketobutyrate were good amino acceptors. We named the enzyme as opine... 

The DPAMP-Rh(I) catalyst system was applied to die asymmetric hydrogenation of methyl (Z)-2-acetamido-3-(3-methoxy-4-acetoxyphenyl)acrylate 147 to furnish the corresponding amino acid 148, which is a crucial advanced intermediate in the synthesis of L-DOPA. Likewise, a homophenylalanine intermediate, which is a key component of the antihypertensive (5,5)-benazepril, was obtained from the hydrogenation of ethyl (Z)-2-acetamido-4-phenylcrotonate using the same catalyst and identical reaction conditions. ... 

The Neber rearrangement has the potential to be a powerfully simplifying transformation in the synthesis of unnatural or isotopically labelled a-amino acids and, indeed, several such syntheses have been reported. Homophenylalanine (44) has been prepared from ethylimidate 42 via chlorination to form the chloroimidate 43. Neber rearrangement of 43 provided 44 in excellent yield, allowing access to labelled versions of this a-amino acid. A synthesis of vinylglycine (47) has also been reported... 

Some nonproteinogenic amino acids, produced enzymatically, are shown in Table 29.4 for example, L-homophenylalanine, a key intermediate of levetiracetam and brivaracetam applicable as antiepileptic drugs, or D-fluoroalanine, a key intermediate of antibiotics inactivating the bacterial D-alanine transaminase. In Table 29.4 a selection of proteinogenic and nonproteinogenic amino acids, the used biocatalysts, synthesis strategies, and the substrates used are listed. 

The asymmetric synthesis via a-transaminases was described for L-phenylalanine and L-homophenylalanine in several reports and reviews [17,62,86]. Herein the focus is on ffl-transaminases that catalyzed the asymmetric synthesis of optically pure nonproteinogenic amino acids as building blocks for peptidomimetic and other pharmaceutical compounds. The overall advantage of -transaminase-catalyzed reactions is the ability to use achiral amino donors like benzylamine, which thermodynamically favors the equilibriinn toward the product side. 

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