Activity and enantioselectivity of the hydroxynitrile lyase MeHNL in dry organic solvents

Abstract

Water concentration affects both the enantioselectivity and activity of enzymes in dry organic media. Its influence has been investigated using the hydrocyanation of benzaldehyde catalyzed by hydroxynitrile lyase cross-linked enzyme aggregate (MeHNL-CLEA) as a model reaction. The enzyme displayed higher enantioselectivity at higher water concentration, thus suggesting a positive effect of enzyme flexibility on selectivity. The activity increased on reducing the solvent water content, but drastic dehydration of the enzyme resulted in a reversible loss of activity.

Scheme 1

MeHNL-catalyzed hydrocyanation of benzaldehyde in different organic solvents.

Figure 1

Variation in ee of (S)-mandelonitrile (3) in the MeHNL-CLEA-catalyzed hydrocyanation in MTBE with different water concentrations (▪ dry; ○ sat.; • 0–2; □ 2–7; ▵ 7–12).

Figure 2

Conversion of 1 in the MeHNL-CLEA-catalyzed synthesis of (S)-mandelonitrile (3) in MTBE with different water concentrations (▪ dry; ○ sat.; • 0–2; □ 2–7; ▵ 7–12).

Figure 3

Effect of water concentration on the conversion of 1 in the MeHNL-CLEA-catalyzed synthesis of (S)-mandelonitrile (3) in MTBE (• 1 h; □ 2 h; ▴ 3 h; ○ 4 h; ▪ 24 h)

Figure 4

Conversion of 1 and ee of (S)-mandelonitrile (3) in the MeHNL-CLEA-catalyzed hydrocyanation in MTBE with and without the salt pair Na₂HPO₄⋅2 H₂O/Na₂HPO₄⋅7 H₂O in the reaction medium (with salt: ▵=conv., ○=ee; without salt: •=conv., ▪=ee).

Figure 5

Variation in ee of (S)-mandelonitrile (3) in the MeHNL-CLEA-catalyzed hydrocyanation in toluene at different water concentrations (▪ dry; ○ sat.; • 0–2; □ 2–7; ▵ 7–12).

Figure 6

Conversion of 1 in the MeHNL-CLEA-catalyzed synthesis of (S)-mandelonitrile (3) in toluene with different water concentrations (▪ dry; ○ sat.; • 0–2; □ 2–7; ▵ 7–12).

Figure 7

Effect of water concentration on the conversion of 1 in the MeHNL-CLEA-catalyzed synthesis of (S)-mandelonitrile (3) in toluene. (• 1 h; ○ 2 h; ▴ 3 h; □ 24 h)

Figure 8

Conversion of 1 in the MeHNL-CLEA-catalyzed synthesis of (S)-mandelonitrile (3) in octane at different water concentrations (▪ dry; ○ sat.; • 0–2; □ 2–7; ▵ 7–12).

Figure 9

TGA of different enzyme preparations in nitrogen at a heating rate of 10 °C min⁻¹.