Ketones enzymic reduction, specificity

Baker s yeast has been widely used for the reduction of ketones. The substrate specificity and enantioselectivity of the carbonyl reductase from baker s yeast, which is known to catalyze the reduction of P-keto ester to L-hydroxyester (L2-enzyme) [15], was investigated, and the enzyme was found to reduce chloro-, acetoxy ketones with high enantioselectivity (Figure 8.32) [24aj. 

Alcohol dehydrogenase-catalyzed reduction of ketones is a convenient method for the production of chiral alcohols. HLAD, the most thoroughly studied enzyme, has a broad substrate specificity and accommodates a variety of substrates (Table 11). It efficiendy reduces all simple four- to nine-membered cycHc ketones and also symmetrical and racemic cis- and trans-decalindiones (167). Asymmetric reduction of aUphatic acycHc ketones (C-4—C-10) (103,104) can be efficiendy achieved by alcohol dehydrogenase isolated from Thermoanaerohium hrockii (TBADH) (168). The enzyme is remarkably stable at temperatures up to 85°C and exhibits high tolerance toward organic solvents. Alcohol dehydrogenases from horse Hver and T. hrockii... 

Zhu, D., Yang, Y., Buynak, J.D. and Hua, L. (2006) Stereoselective ketone reduction by a carbonyl reductase from Sporobolomyces salmonicolor. Substrate specificity, enantioselectivity and enzyme—substrate docking studies. Organic and Biomolecular Chemistry, 4 (14), 2690-2695. 

Using two types of specially synthesized rhodium-complexes (12a/12b), pyruvate is chemically hydrogenated to produce racemic lactate. Within the mixture, both a d- and L-specific lactate dehydrogenase (d-/l-LDH) are co-immobilized, which oxidize the lactate back to pyruvate while reducing NAD+ to NADH (Scheme 43.4). The reduced cofactor is then used by the producing enzyme (ADH from horse liver, HL-ADH), to reduce a ketone to an alcohol. Two examples have been examined. The first example is the reduction of cyclohexanone to cyclohexanol, which proceeded to 100% conversion after 8 days, resulting in total TONs (TTNs) of 1500 for the Rh-complexes 12 and 50 for NAD. The second example concerns the reduction of ( )-2-norbornanone to 72% endo-norbor-nanol (38% ee) and 28% exo-norbornanol (>99% ee), which was also completed in 8 days, and resulted in the same TTNs as for the first case. 

Aldehyde reductases are a group of isoenzymes that catalyze the NADPH-specific reduction of aldehydes. Ketones do not serve as substrates for these enzymes. The best substrates for aldehyde reductase are aromatic aldehydes and those aldehydes obtained through metabolism of biogenic amines. The species distribution, specificity, and inhibition of aldehyde reductases have been reviewed (792). 

Alcohol dehydrogenase-catalyzed reduction of ketones is a convenient method lor the production nf chiral alcohols. HI.AD, the most thoroughly studied enzyme, has a broad substrate specificity and accommodates a variety of substrates. 

Reactions with these compounds suffer from very low substrate concentrations due to the low solubility of hydrophobic ketone substrates in aqueous media, which leads to unsatisfactory volumetric productivities. To achieve higher substrate concentrations, a biphasic reaction medium was introduced. The system water/ n-heptane (4 1) proved to be the most suitable system with regard to stability of the examined enzymes. The large-scale available (S)-specific ADH from R. erythropolis as well as FDH from C. boidinii are stable for long periods of time in this aqueous-organic solvent system. Preparative conversions with a variety of aromatic ketone substrates were carried out with this reaction medium. For example, p-chloroacetophenone was converted into the corresponding (S )-alcohol with >99% ee and 69% conversion. The obvious increase in volumetric productivity is due to the higher substrate concentrations. The reduction of p-chloroacetophenone... 

However from the standpoint of green chemistry, the use of isolated enzymes (or dead whole cells) is highly preferred because it avoids the generation of copious amounts of biomass. It must be emphasized that the productivity of microbial conversions is usually low, since non-natural substrates are only tolerated at concentrations as low as 0.1-0.3% [106]. The large amount of biomass present in the reaction medium causes low overall yields and makes product recovery troublesome. Therefore the E-factors for whole cell processes can be extremely high. Moreover the use of wild-type cells often causes problems because an array of enzymes is present which can interfere in the reduction of a specific ketone (giving opposite selectivities). The use of recombinant techniques, however, which only express the desired enzyme can overcome this problem [108]. 

Reductions of symmetrical bicyclic diketones may also be effected selectively, as illustrated in Scheme 12. The stereospecificities of the HLADH-catalyzed transformations of the unsaturated decalin-diones (25) and (27) to the corresponding hydroxy ketones (26) and (28), and in fact all specificity aspects of this enzyme, are fully predictable using a simple to use, cubic-space model of the enzyme s... 

Regiospeciflcity combined with diastereotopic face specificity is seen in the reduction of the trione (60) to (61) (Scheme 26). ° In Scheme 27 the reduction is quite regiospeciflc, but both diastereotopic feces of the exocyclic ketone function of (62) are attacked to give the erythro-(63) and threo- 64) hydroxy ketone products. The degree to which regio specificity and diastereotopic face specificity can be controlled with different enzymes is shown in the bile acid reductions of Scheme 28. ° ... 

Enantiomeric and diastereotopic face specific reductions are also readily effected on racemic bicyclic ketones. An illustration of the broad structural range that is amenable to enzyme-catalyzed transformation in this way is given in Scheme 38. While 2-decalones, such as ( )-(81)- 83), and the related heterocyclic analogs ( )-(85) are good substrates for HLADH, the 1-decalone ( )-(84) is not. However, by changing enzymes to MJADH, ( )-(84) becomes a good substrate.Similarly, TBADH is a highly satisfactory catalyst for stereospecific reduction of ( )-(86), but will not accept its dimethyl... 

Enzyme-mediated carbonyl reductions are not restricted to aldehydes and ketones. With some organisms, carboxy groups can be reduced to primary alcohols. The example in Scheme 53 proceeds with enantiomeric specificity, ... 

Unlike aldehydes, ketones are essentially unreactive, apart from being reduced when the equilibrium shifts predominantly to the secondary alcohol that is readily conjugated, thereby encouraging further reduction of the ketone [Eq. (14)]. If ketone reduction results in the formation of an asymmetric carbon, a high degree of stereospecificity often occurs, even though the enzyme systems demonstrate a low degree of substrate specificity. 

The cubic section models, co13 and co065, proved to be accurate in predicting acceptance of substrates and the stereochemical course of HLADH mediated reductions. The average priority value for all cubes, number of forbidden sites and number of low priority cubes occupied by atoms in a possible alcohol are used to estimate the acceptability of the corresponding ketone by HLADH. The prediction method developed can be applied to other enzymes (e.g. other dehydrogenases, esterases, or lipases) if kinetic and specificity data is available. 

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