Epoxide hydrolases resolutions

Fig. 17. Bacterial epoxide hydrolase. Resolution of ( )-l-methyl-l,2-epoxycyclohexane [192] and ( )-l,2-epoxyoctane [193] using respectively whole cells of Corynebacterium C12 and Chryseomonas luteola...

Epoxide hydrolases Resolution of aryl and substituted alicyclic epoxides. Enantioselective catalyst... 

Several reports regarding the directed evolution of enantioselective epoxide hydrolases (EHs) have appeared [23,57-59]. These enzymes constitute important catalysts in synthetic organic chemistry [4,60]. The first two reported studies concern the Aspergillus niger epoxide hydrolase (ANEH) [57,58]. Initial attempts were made to enhance the enantioselectivity of the AN E H -catalyzed hydrolytic kinetic resolution of glycidyl phenyl ether (rac-19). The WT leads to an Evalue of only 4.6 in favor of (S)-20 (see Scheme 2.4) [58]. 

Figure 2.15 Iterative CASTing in the evolution of enantioseiective epoxide hydrolases as catalysts in the hydrolytic kinetic resolution ofrac-19[23].

Lewis DF, Lake BG, Bird MG. Molecular modelling of human microsomal epoxide hydrolase (EH) by homology with a fungal (Aspergillus niger) EH crystal structure of 1.8 A resolution structure-activity relationships in epoxides inhibiting EH activity. Toxicol In Vitro 2005 19 517-22. 

J. Zou, B. M. Hallberg, T. Bergfors, F. Oesch, M. Arand S. L. Mowbray, T. A. Jones, Structure of Aspergillus tiiger Epoxide Hydrolase at 1. 8 A Resolution Implications for the Structure and Function of the Mammalian Microsomal Class of Epoxide Hydrolases , Structure 2000, 8, 111 - 122. 

The use of enzymes and whole cells as catalysts in organic chemistry is described. Emphasis is put on the chemical reactions and the importance of providing enantiopure synthons. In particular kinetics of resolution is in focus. Among the topics covered are enzyme classification, structure and mechanism of action of enzymes. Examples are given on the use of hydrolytic enzymes such as esterases, proteases, lipases, epoxide hydrolases, acylases and amidases both in aqueous and low-water media. Reductions and oxidations are treated both using whole cells and pure enzymes. Moreover, use of enzymes in sngar chemistiy and to prodnce amino acids and peptides are discnssed. 

Figure 2.18 Resolutions of epoxides using microsomal epoxide hydrolases.

Generally monosubstituted and dx-l,2-disubstituted epoxides are good substrates for EH while tri-, tetra or tra s-l,2-disubstituted ones are poor or non-substrates. Resolutions of epoxides using microsomal epoxide hydrolases, mEHs show that cis-2-alkyl substituted styrene oxides gave very high E-values when R=Me or Et (Figure 2.18a). A series of cis-... 

Figure 27. Epoxide hydrolase catalyzed kinetic resolution of c/.v-2-ethyl-3-methyloxirane and formation of 2i ,3f -2,3-pentanediol as monitored by complexation gas chromatography on 0.08 M nickel(II) bis[3-(heptafluorobutanoyl)-(1 / )-camphorate] in methylpolysiloxane [25 m x 0.25 mm (i.d.) glass capillary column. 95CC, 1.1 bar nitrogen]191 2,3-pentanediol as acetonides 0.14 M nickel(ll) bis[3-(heptafluo-robutanoyl)-(l /t,2S)-pinan-4-onatc]151 in SE-30. Note that there is a change in the numbering of the chiral carbon atoms of the oxiranc vs. the diol due to nomenclature requirements.

Epoxide hydrolase Kinetic resolution of epoxides in Tris buffer/ octane [18]... 

The MS assay has also been applied successfully in the directed evolution of enantioselective epoxide hydrolases acting as catalysts in the kinetic resolution of chiral epoxides [35]. Moreover, Diversa has recently employed the MS-based technique for desymmetrization of a prochiral dinitrile catalyzed by mutant nitrilases [36]. In this industrial application one of the nitrile moieties was labeled with 15N, which means that the two pseudo enantiomeric products differ by only one mass unit. 

Figure 18.22. Dynamic kinetic resolution of 2,2-disubstituted epoxides 63 by epoxide hydrolase.

Monfort, N., Archelas, A., and Furstoss, R. 2004. Enzymatic transformations. Part 55 Highly productive epoxide hydrolase catalyzed resolution of an azole antifungal key synthon. Tetrahedron, 60(3), 601-605. 

Zha D, Wilensek S, Hermes M, Jaeger K-E, Reetz MT (2001) Complete reversal of enantioselectivity of an enzyme-catalyzed reaction by directed evolution. Chem Commun (Cambridge UK) 2664-2665 Zou JY, Hallberg BM, Bergfors T, Oesch F, Arand M, Mowbray SL, Jones TA (2000) Structure of Aspergillus niger epoxide hydrolase at 1.8 resolution Implications for the structure and function of the mammalian microsomal class of epoxide hydrolases. Structure (London) 8 111-122... 

Several specific cases have been described in the literature [10,50], A new example concerns the hydrolytic kinetic resolution of the epoxide 1 catalyzed by an epoxide hydrolase (Fig. 11.5) [52], As in other kinetic resolutions, the reaction is allowed to reach the ideal value of 50 %. Instead of employing a genuine racemate (R)-1/(S)-1... 

The combination of a large number of chemical resolution and assignment studies (Table 3) of benzo-ring /rans-dihydrodiols along with studies designed to define the stereochemical course of metabolism of benzo[a]pyrene, ° benz[a]-anthracene, chrysene, phenanthrene, benzo[c]phenanthrene, anthracene, and naphthalene has led to the conclusion that the combined action of cytochrome P450c and epoxide hydrolase has a marked preference for the formation of (—)-(R,R)-dihydrodiols. This result pertains even when the initially... 

In biocatalysis, hydrolases are the most important class of enzymes for carrying out enzymatic resolutions. Many hydrolases, such as esterases, lipases, epoxide hydrolases, proteases, peptidases, acylases, and amidases, are commercially available a substantial number of them are bulk enzymes [87]. Resting-cell systems, if they are not immobilized, are used in diluted suspensions and could be handled as quasi-homogeneous catalysts. 

Resolution of Racemic Epoxides Using Epoxide Hydrolases. 176... 

Fig. 20. Resolution of styrene oxide by A. niger and B. sulfurescens epoxide hydrolases. Deracemization of styrene oxide by using a mixture of these two fungi [175]...

Fig. 23. Fungal epoxide hydrolase. Preparative scale resolution of para-bromo styrene oxide using a two-liquid-phase process [213]...

The presence of epoxide hydrolase activities of fungal cells has been described by several authors [196-201]. However, the real breakthrough in the use of such enzymes for preparative scale synthesis was developed by the group of Furstoss et al. One of their first results was the description of the efficient resolution of racemic geraniol AT-phenylcarbamate by a culture of the fungus A. niger, leading... 

Scheme 2.15 gives some examples of the use of epoxide hydrolases in organic synthesis. Entries 1 to 3 are kinetic resolutions. Note that in Entry 1 the hydrolytic product is obtained in high e.e., whereas in Entry 2 it is the epoxide that has the highest e.e. In the first case, the reaction was stopped at 18% conversion, whereas in the second case hydrolysis was carried to 70% completion. The example in Entry 3 has a very high E (> 100) and both the unreacted epoxide and diol are obtained with high e.e. at 50% conversion. Entry 4 shows successive use of two separate EH reactions having complementary enantioselectivity to achieve nearly complete... 

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