Asymmetric Retro‐Henry Reaction Catalyzed by Hydroxynitrile Lyase from Hevea brasiliensis

Abstract

Hydroxynitrile lyase from Hevea brasiliensis (HbHNL) is a promiscuous biocatalyst that, besides the native cyanohydrin reaction, also catalyzes the asymmetric Henry reaction yielding (S)‐β‐nitroalcohols with high enantiomeric excess. Since the Henry reaction is reversible, the enzyme can be also utilized for the production (R)‐enantiomers by means of resolution of racemic β‐nitroalcohols. Herein the biocatalytic retro‐Henry reaction is studied using the cleavage of 2‐nitro‐1‐phenylethanol as a model system. The main problem that prevents high levels of conversion or high ee values during the cleavage of the β‐nitroalcohol is the formation of benzaldehyde, which is known to be a strong enzyme inhibitor. The product inhibition is overcome by performing the biocatalytic retro‐Henry reaction in the presence of HCN, which reacts in situ with benzaldehyde and converts it to the less‐inhibitive mandelonitrile. By using such a reaction cascade, it was possible to conduct the resolution practically to completion (95 % ee, 49 % conversion). Furthermore, the catalyst productivity achieved during the resolution was ten times higher than that in the HbHNL‐catalyzed synthesis of (S)‐2‐nitro‐1‐phenylethanol by condensation of benzaldehyde and nitromethane.

Synthesis of (R)-β-nitro alcohols catalyzed by R-selective hydroxynitrile lyase from Arabidopsis thaliana in the aqueous–organic biphasic system

“…Only recently, a biocatalytic asymmetric reaction using HbHNL that affords (S)-␤-nitro alcohols with high enantiomeric excess (>99%) has been reported (Gruber-Khadjawi et al, 2007;Purkarthofer et al, 2006). Although only the (S)-␤-nitro alcohols can be produced in this manner, the reversibility of Henry reaction enables in principle the application of HbHNL also the production of complementary (R)-␤-nitro alcohols by means of kinetic resolution not asymmetric synthesis (Yuryev et al, 2010).…”

Increasing the Reaction Rate of Hydroxynitrile Lyase fromHevea brasiliensistoward Mandelonitrile by Copying Active Site Residues from an Esterase that Accepts Aromatic Esters

“…rac -2-Nitro-1-phenylethanol was synthesized according to a literature method. [25] Assay conditions: (190-x) µL 50 mM citrate buffer pH 5.0,×µL 100 mM stock solution of rac-mandelonitrile (HNL) or rac -2-nitro-1-phenylethanol (nitroaldolase) in 10 mM citric acid to suppress spontaneous cleavage and 10 µL enzyme sample (200 µL total volume within a microtiter plate; path length of 0.6 cm). If required, up to 50% acetonitrile can be added to the substrate stock solution to dissolve the substrate.…”

Biocatalytic Approach to Chiral β-Nitroalcohols by Enantioselective Alcohol Dehydrogenase-Mediated Reduction of α-Nitroketones

“…Several examples of kinetic resolution of racemic compounds 1 catalysed by hydrolases have been reported in the literature [7]. It has also been discovered that some hydroxynitrile 2 of 11 lyases (HNLs) are able to promote the enantioselective addition of nitromethane to aldehydes, such as the (S)-selective HNLs from Hevea brasiliensis and from Manihot esculenta [18][19][20], and the (R)-selective HNLs from Arabidopsis thaliana [21], Acidobacterium capsulatum, and Granulicella tundricula [22]. These reactions are generally characterized by long reaction times, and strong substrate dependence.…”

“…[18][19][20], and the (R)-selective HNLs from Arabidopsis thaliana [21], Acidobacterium capsulatum, and Granulicella tundricula [22]. These reactions are generally characterized by long reaction times, and strong substrate dependence.…”