Enzymatics stereospecificity

Hill, R. K. Enzymatic Stereospecificity at Prochiral Centers of Amino Acids, in Bio-organic chemistry, Vol. 2, p. 111. (van Tamelen, E. E. ed.). New York Academic Press 1978... 

The second task, resolution of synthetic D,L-amino acids has been solved by conversion of the neutral amino acids into real carboxyUc acids by acylation of the amino group, either by the benzoyl or the formyl residue. The D,L-acyl amino acids then formed diastereomeric salts with optically active bases, mostly alkaloids, which differed in their solubihty in various solvents, and so could be separated by recrystallization. This method is still in use, although enzymatic procedures, specific oxidation of the D-antipode in the presence of D-amino acid oxidase or enzymatic, stereospecific removal of iV-acyl residues from d,l-AT-acetyl-amino acids by acylase, are more convenient. Certainly, L-amino acids became accessible from nature by the ester method, but without synthetic material, extended experiments of peptide couphng would have been impossible. 

Most of the enzymatic peptide forming reactions are strictly stereo-specific for L-amino acids so that racemization that often accompanies chemical coupUng of optically active amino acids does not occur. The formation of only the L-amino acid derivative out of a D,L-mixture, in an enzymatic formation e.g. of an anilide from a D,L-Z-amino acid ester and aniUne, makes proteolytic enzymes useful reagents for the resolution of racemic mixtures. This is supplementary to the enzymatic stereospecific deacylation of D,L-iV-acylamino acid mixtures where exclusively the L-derivative will be deacylated. 

Other enzymes widely used in organic syntheses are lipases. Lipases are serine hydrolases that catalyze the hydrolysis of lipids to fatty acids. An example of their use is in the resolution of racemic methyl 4-benzyloxy-2-(hydroxymethyl)[l- C]butyrate (8), an intermediate for the carbon-13 labeling of the antiviral agents penciclovir and famciclovir. From a limited number of candidates Candida cylindracea lipase turned out to be the enzyme of choice. At optimal reaction conditions (pH 5-7, 35 °C) no nonenzymatic hydrolysis was detectable. Due to insufficient enzymatic stereospecificity, however, the hydrolysis provided the unreacted dextrorotary ester 2 at an optical purity of 94% e.e. and the hydrolyzed levorotary acid 10 at an optical purity of 70% e.e. 

In many cases only the racemic mixtures of a-amino acids can be obtained through chemical synthesis. Therefore, optical resolution (42) is indispensable to get the optically active L- or D-forms in the production of expensive or uncommon amino acids. The optical resolution of amino acids can be done in two general ways physical or chemical methods which apply the stereospecific properties of amino acids, and biological or enzymatic methods which are based on the characteristic behavior of amino acids in living cells in the presence of enzymes. 

Af-(ter -Butoxycarbonyl)-3-hydroxymethylpiperidine is a building block in the synthesis of a potent tryptase inhibitor. It is produced by stereospecific esterification of the racemic alcohol with succinic anhydride [30]. The enzymatic resolution is followed up by separation of... 

Kanegafuchi Chemical Industries produce D-p-hydroxyphenyl glycine, which is a key raw material for the semisynthetic penicillins ampicillin and amoxycillin. Here, an enantioselective hydantoinase is applied to convert the hydantoin to the D-p-hydroxyphenyl glycine. The quantitative conversion of the amide hydrolysis is achieved because of the in situ racemization of the unreacted hydantoins. Under the conditions of enzymatic hydrolysis, the starting material readily racemizes. Therefore, this process enables the stereospecific preparation of various amino acids at a conversion of 100% [38]. 

It was shown that microsomal epoxide hydrolase-catalyzed trans-addition of water to BaP 9,10-epoxide occurs stereospecifically at the C-9 position (15). Since BaP is metabolized essentially to an optically pure 9R,10R-dihydrodiol (13 and L5 Table I), the 9,10-epoxide formed in BaP metabolism must have 9S,10R absolute stereochemistry (Figure 1). Similarly, the 7,8-epoxide formed in BaP metabolism is hydrated specifically at the C-8 position to form the 7R,8R-dihydrodiol (14.21). Hence the enzymatically formed 7,8-epoxide intermediate has 7R,8S absolute stereochemistry (Figure 1). Although the 7R,8R-dihydrodiol is formed almost exclusively from BaP metabolism in rat liver microsomes (Table I) and in bovine bronchial explants (25). the 7S,8S-dihydrodiol is also formed from BaP metabolism in mouse skin epidermis in vivo (5). 

On the other hand a stereospecific reaction is one in which a particular stereoisomeric form of the reactant reacts to give a stereoisomeric form of the product. The enzymatic reduction of pyruvic acid is not stereospecific because the reactant can not existin stereoisomeric form. Therefore, all reactions that are stereospecific are necessarily stereoselective but not all stereoselective reactions are stereospecific. 

All evidence presented here indicates that the methoxime-to-ketone reaction is enzymatic, but the nature of the reaction is unclear. Hydrolysis by an uncharacterized hydrolase, as postulated by the authors, is a valid assumption. However, one should not dismiss another possibility, namely oxidative O-demethylation to the oxime, followed by spontaneous oxime-to-ketone hydrolysis, and stereospecific reduction, as explained above. 

While the uncatalyzed reaction requires extremely high temperatures and proceeds nonspecifically, the enzymatic reaction proceeds regiospecifically and stereospecifically under physiological conditions of temperature and pressure. The P450 enzyme utilizes its iron-porphyrin centers as metal-containing... 

Reactions catalyzed by enzymes or enzyme systems exhibit far greater specificities than more conventional organic reactions. Among these specificities which enzymatic reactions possess, stereospecificity is one of the most excellent. To overcome the disadvantage of a conventional synthetic process, i.e., the troublesome resolution of a racemic mixture, microbial transformation with enzymes possessing stereospecificities has been appHed to the asymmetric synthesis of optically active substances [1-10]. C3- and C4-synthetic units (synthons, building blocks), such as epichlorohydrin (EP), 2,3-dichloro-l-propanol (2,3-DCP), glycidol (GLD), 3-chloro-l,2-propanediol (3-CPD), 4-chloro-... 

Based on the stereospecific transketolase-catalyzed ketol transfer from hydroxy-pyruvate (20) to D-glyceraldehyde 3-phosphate (18), we have thus developed a practical and efficient one-pot procedure for the preparation of the valuable keto-sugar 19 on a gram scale in 82% overall yield [29]. Retro-aldolization of D-fructose 1,6-bisphosphate (2) in the presence of FruA with enzymatic equilibration of the C3 fragments is used as a convenient in-situ source of the triose phosphate 18 (Scheme 2.2.5.8). Spontaneous release of CO2 from the ketol donor 20 renders the overall synthetic reaction irreversible [29]. 

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