Enzymatic synthesis applications

Enzymes are generally classified into six groups. Table 1 shows typical polymers produced with catalysis by respective enzymes. The target macromolecules for the enzymatic polymerization have been polysaccharides, poly(amino acid)s, polyesters, polycarbonates, phenolic polymers, poly(aniline)s, vinyl polymers, etc. In the standpoint of potential industrial applications, this chapter deals with recent topics on enzymatic synthesis of polyesters and phenolic polymers by using enzymes as catalyst. 

In polyester synthesis via ring-opening polymerizations, metal catalysts are often used. For medical applications of polyesters, however, there has been concern about harmful effects of the metallic residues. Enzymatic synthesis of a metal-free polyester was demonstrated by the polymerization of l,4-dioxan-2-one using Candida antarctica lipase (lipase CA). Under appropriate reaction conditions, the high molecular weight polymer (molecular weight = 4.1 x 10" ) was obtained. 

Ritter H (1996) Functionalized polymers via enzymatic synthesis. In Arshady R (ed) Desk references of functional polymers syntheses and applications. ACS, Washington,... 

S. Kojima, T. Hasegawa, T. Yonemura, K. Sasaki, K. Yamamoto, Y. Makimura, T. Takahashi, T. Suzuki, Y. Susuki, and K. Kobayashi, Ruthenium complexes carrying a disialo complex-type oligosaccharide Enzymatic synthesis and its application to a luminescent probe to detect influenza viruses, Chem. Commun. (2003) 1250-1251. 

The third contribution presents the combination of electrochemistry and enzymatic synthesis for the selective formation of complex molecules. This quite young field of research is developing rapidly because the application of the reagent-free electrochemical procedure combined with the regio- and stereoselectivity of enzymes offers the possibility of establishing new environmentally friendly process even on a technical scale. 

Other mediators which have been used in combination with diaphorase for the regeneration of NAD+ are riboflavin and Vitamin K3, which is 2,3-dimethyl-1,4-naphthoquinone. However, especially riboflavin is not stable enough for synthetic applications [40]. Better stability is exhibited by phenanthrolindiones as mediators. In combination with diaphorase, Ohshiro [41] showed the indirect electrochemical oxidation of cyclohexanol to cyclohexanone using the NAD+ dependent HLADH with a turnover frequency of two per hour. For an effective enzymatic synthesis, this turnover frequency, however, would be too small. In our own studies, we were able to accelerate the NAD(P)+ regeneration considerably by lowering the electron density within the... 

The procedure can provide a higher amount of L-pipecolic acid in a shorter reaction time than the previously reported system, indicating that it is applicable in industrial production of L-pipecolic acid. A similar system was successfully employed in the enzymatic synthesis of several cyclic amino acids by our group. ... 

Reetz et al. described the solid-phase enzymatic synthesis of oligonucleotides on Kieselguhr-PDMA-resins via T4 RNA ligase. Goncomitantly, they found that RNase A selectively cleaves the last bound nucleotide at the ribose sugar leaving a 3, 5 - diphosphorylated ohgomer on the resin, but application in synthesis has not yet been undertaken [22]. 

Rare or unnatural monosaccharides have many useful applications as nonnutritive sweeteners, glycosidase inhibitors and so on. For example, L-glucose and L-fructose are known to be low-calorie sweeteners. In addition, rare or unnatural monosaccharides are potentially useful as chiral building blocks for the synthesis of biologically active compounds. Therefore, these compounds have been important targets for the development of enzymatic synthesis based in the use of DHAP-dependent aldolases alone or in combination with isomerases. Fessner et al. showed that rare ketose-1-phosphates could be reached not only by aldol addition catalyzed by DHAP-dependent aldolases, but by enzymatic isomerization/ phosphorylation of aldoses [35]. Thus, for example, L-fructose can be prepared... 

Catalytic Oxidation of NAD(P)H A Continuously Improved Selection of Suitable ROMs This research is triggered hy at least two reasons (1) the importance of NAD(P)H/NAD(P)- - redox couples in biological systems is known, as is known the dependence of oxidation mechanisms on the oxidants [14, 82, 172-174] (2) the possibility of developing amperometric biosensors for NAD(P)+-dependent dehydrogenases. As a consequence, much attention is devoted to the regeneration of these coenzymes in their reduced or oxidized forms for their application in biosensors or in enzymatic synthesis [180]. Here, we are concerned with electrochemical regeneration [181]. 

The application of SCF as reaction media for enzymatic synthesis has several advantages, such as the higher initial reaction rates, higher conversion, possible separation of products from unreacted substrates, over solvent-free, or solvent systems (where either water or organic solvents are used). Owing to the lower mass-transfer limitations and mild (temperature) reaction conditions, at first the reactions which were performed in non-aqueous systems will be transposed to supercritical media. An additional benefit of using SCFs as... 

Relatively few of the enzymatic methods applicable to the preparation of secondary cyanohydrins have been adapted successfully to the synthesis of optically pure tertiary cyanohydrins [1,3,4]. Similarly, progress in asymmetric hydro-cyanation of ketones with synthetic catalysts lagged far behind advances in aldehyde cyanation. This situation has changed fairly dramatically over the past... 

More effort was therefore invested in the application of synthetic methodologies for these alkaloids and some straightforward chemo-enzymatic approaches were recently developed [150]. An enzyme-catalysed aldol reaction was again a crucial step in that synthetic route and is strongly reminiscent of Wong s research strategy relating to the chemo-enzymatic synthesis of pyrrolizidines mentioned earlier. 

Chen, S-T- Fang, J-M. Preparation of optically active tertiary alcohols by enzymatic methods. Application to the synthesis of drags and natural products. J. Org. Chem. 1997, 62, 4349-4357. 

The enzymatic synthesis of glycopeptides does not require protection of the functional groups of the amino acid side chains and sugar hydroxyls, because of the high stereoselectivity and regioselectivity of proteases. However, the substrate selectivity of these enzymes may limit a wider range of applications. 

Petersson, A.E.V., L.M. Gustafsson, M. Nordblad, P Borjesson, B. Mattiasson and P. Adlercreutz, Wax Esters Produced by Solvent-Free Energy-Efficient Enzymatic Synthesis and Their Applicability as Wood Coatings, Green Chemistry, 7, 837-843 (2005). 

The number of enzymes for industrial synthetic applications is growing fast. Enzymatic synthesis can be performed under mild reaction conditions so that many problems of chemical synthesis like isomerization orracemization can be prevented. Furthermore, enzymes are highly specific and selective, especially for enantio- or regio-selective introduction of functional groups. For the preparation of chiral enantiopure compounds, the resolution of racemic mixtures by hydrolases is a well-established route, which has the advantage to be able to use enzymes free of coenzymes. Otherwise, only a maximum yield of 50% can be reached by the primary reaction and further steps of reracemization must follow to avoid loss of the undesired enantiomer. 

This article reviews the main applications of glycosidases and lipases in the synthesis of glycosidic bonds and in acylation/deacylation reactions of carbohydrates, respectively. Special attention is given to the factors that can affect the selectivity of the reactions, such as the enzyme origin, the structure of the substrates, and the reaction medium. A number of reviews have appeared in the literature on enzymatic synthesis of carbohydrates that include reactions with glycosidases and/or lipases [ 1 - 6]. In this article only a selection of examples are given to illustrate the discussion, rather than an extensive compilation of the work published in the area. 

Different types of antitemplates can be and, presumably, will be, synthesized by employing other chemical reactions that are sufficiently mild and selective for the modification of polynucleotides. Another approach to this problem is, of course, to synthesize an appropriately modified nucleoside triphosphate and utilize it as one of the substrates in a DNA or RNA polymerizing system, in the presence of the specific template for which an antitemplate is to be made. The enzymatic synthesis method has some obvious advantages however, the inhibitory activity of the product (antitemplate) may limit its applications. 

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