Optimization of arylacetonitrilase production from Alcaligenes sp. MTCC 10675 and its application in mandelic acid synthesis

Abstract

Alcaligenes sp. MTCC 10675 has been isolated from soil sample using enrichment method and has nitrilase catalytic system which is highly specific for the hydrolysis of arylaliphatic nitriles. Optimization of culture conditions using response surface methodology and inducer-mediated approach enhanced arylacetonitrilase production significantly (2.4-fold). Isobutyronitrile acted as an effective inducer for the induction of arylacetonitrilase, and it is highly specific for arylacetonitriles (phenyl acetonitrile and mandelonitrile). Arylacetonitrilase has no effect on its relative velocity (V r) up to 20 mM substrate (mandelonitrile) concentration and at 30 mM mandelonitrile, 23.4 % degree of inhibition (I d) was recorded. Half life of arylacetonitrilase of Alcaligenes sp. MTCC 10675 was 27.5 h at 25 °C. Hg(2+), Ag(+), Pb(3+), and Co(2+) were strong inhibitor of arylacetonitrilase activity which resulted into 100 %, 91 %, 84 %, and 83 % inhibition, respectively. Polar protic solvent (dichloromethane, dimethylsulphooxide, and n-butanol) reduce arylacetonitrilase activity up to 80-94 % at 10 % concentration. Alcaligenes sp. MTCC 10675 has higher biocatalytic activity, i.e., 3.9 gg(-1) dcw, which is highest in comparison to till reported organism. Arylacetonitrilase-mediated hydrolysis of racemic mandelonitrile resulted into R-(-) mandelic acid with 99.0 % enantiomeric excess (e.e.).

Purification and characterization of arylacetonitrile‐specific nitrilase of Alcaligenes sp. MTCC 10675

“…Alcaligenes sp. MTCC 10675 (isolated from soils of Shimla District of Himachal Pradesh, India, as a nitrile‐metabolizing bacterium and identified at the Microbial Type Culture Collection and Gene Bank, Institute of Microbial Technology, Chandigarh, India, was grown overnight in a seed medium (nutrient broth), and 1 mL of the seed culture was transferred into a 250 mL Erlenmeyer flask containing 50 mL of production medium (pH 7.0) consisting of (g/L) peptone 5.5 g, malt extract 3.0 g, yeast extract 5.5 g, and isobutyronitrile 4.0 g (60 mM) at 30 °C for 24 H . After 24 H, cells were separated from the culture broth by centrifugation at 10,000 g , washed twice with 0.1 M K 2 HPO 4 /KH 2 PO 4 (pH 7.0), and suspended in the same buffer containing 1 mM dithiothreitol (DTT) and 1 mM ethylenediaminetetraacetate (EDTA).…”

“…A nitrile‐degrading bacterium Alcaligenes sp. MTCC 10675 has been isolated in our laboratory, which has high affinity for arylacetonitriles . In this article, purification, characterization, and sequence analysis of the amino acid sequence of arylacetonitrile‐hydrolyzing nitrilase of Alcaligenes sp.…”

Synthesis and Characterization of Chiral Iridium Complexes Bearing Carbohydrate Functionalized Pyridincarboxamide Ligands and Their Application as Catalysts in the Asymmetric Transfer Hydrogenation of α-Ketoacids in Water

“…In this context, the hydrogenation of CO double bonds of α-ketoesters and α-ketoacids has a special importance, because the corresponding chiral α-hydroxyacids stand out for their synthetic and biological relevance (Scheme ). − For example, ( R )-(−)-mandelic acid (R = Ph) is an important pharmaceutical ingredient, whereas both enantiomers of 3-phenyllactic acid (R = CH 2 Ph) are antimicrobial agents active against bacteria and fungi, and the same synthons are claimed as monomers of bioderived aromatic polymers. , …”

Classifying nitrilases as aliphatic and aromatic using machine learning technique

“…Nitrilases differ variably in substrate specificities and find wide application in the transformation of a range of nitriles to acids (Sharma et al 2006(Sharma et al , 2012Bhatia et al 2014). Previous studies have revealed that nitrilases are specific for aromatic nitriles while nitrile hydratase has affinity towards aliphatic nitriles, but in light of rapidly growing information regarding nitrile metabolizing enzymes, various aspects have to be reconsidered (Mylerova and Martinkova 2003).…”