Nitrile hydratase of Rhodococcus erythropolis: characterization of the enzyme and the use of whole cells for biotransformation of nitriles

The intracellular cobalt-type nitrile hydratase was purified from the bacterium Rhodococcuserythropolis. The pure enzyme consisted of two subunits of 29 and 30 kDa. The molecular weight of the native enzyme was estimated to be 65 kDa. At 25 °C the enzyme had a half-life of 25 h. The Michaelis–Menten constants K(m) and v(max) for the enzyme were 0.624 mM and 5.12 μmol/min/mg, respectively, using 3-cyanopyridine as the substrate. The enzyme-containing freely-suspended bacterial cells and the cells immobilized within alginate beads were evaluated for converting the various nitriles to amides. In a packed bed reactor, alginate beads (2 % alginate; 3 mm bead diameter) containing 200 mg/mL of cells, achieved a conversion of >90 % for benzonitrile and 4-cyanopyridine in 38 h (25 °C, pH 7.0) at a feed substrate concentration of 100 mM. The beads could be reused for up to six reaction cycles

Demonstration of Redox Potential of Metschnikowia koreensis for Stereoinversion of Secondary Alcohols/1,2-Diols

The present work reports the Metschnikowia koreensis-catalyzed one-pot deracemization of secondary alcohols/1,2-diols and their derivatives with in vivo cofactor regeneration. Reaction is stereoselective and proceeds with sequential oxidation of (R)-secondary alcohols to the corresponding ketones and the reduction of the ketones to (S)-secondary alcohols. Method is applicable to a repertoire of racemic aryl secondary alcohols and 1,2-diols establishing a wide range of substrate specificity of M. koreensis. This ecofriendly method afforded the product in high yield (88%) and excellent optical purity (>98% ee), minimizing the requirement of multistep reaction and expensive cofactor

Efficient chemoenzymatic synthesis of (RS)-, (R)-, and (S)-bunitrolol

A new chemical and the first chemoenzymatic synthesis of β-adrenergic receptor blocking agent bunitrolol is reported in racemic (RS) and enantioenriched forms (R and S). The intermediates (R)- and (S)-1-chloro-3-(2-cyanophenoxy)propan-2-ol intermediates were synthesized from the corresponding racemic alcohol through enzymatic kinetic resolution. The commercial available lipases PS-C and CCL exhibited complementary enantioselectivity during transesterification of the racemic alcohol with vinyl acetate affording the (R)-alcohol along with (S)-acetate and the (S)-alcohol along with (R)-acetate, respectively, and represent an example of enzymatic switch for reversal of enantioselectivity. The effects of various reaction parameters, such as temperature, time, substrate and enzyme concentration, and reaction medium, on the activity and enantioselectivity were optimized. The (R)- and (S)-alcohols were converted into (S)-and and (R)-bunitrolol, respectively, by treatment with tert-butylamine. The (R)- and (S)-acetates, obtained enzymatically were deacetylated to the corresponding alcohol by chemical hydrolysis and further converted into (S)-and and (R)-bunitrolol by chemical means. This is the first chemoenzymatic synthesis of both of the enantiomers of the drug. (RS)-, (R)-, and (S)-Bunitrolol were also synthesized following the ‘all chemical’ routes from (RS)-, (R)-, and (S)-epichlorohydrin via the corresponding (RS)-, (S)-, and (R)-2-cyanoglycidyl ether and the (RS)-, (R)-, and (S)-1-chloro-3-(2-cyanophenoxy)propan-2-ol intermediates with improved overall yields and better enantiomeric excesses compared to the reported processes