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

Abstract

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 Km and vmax 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.

Evaluation of acrylamide production by Rhodococcus rhodochrous (RS-6) cells immobilized in agar matrix

“…At the end of each cycle, the reaction was terminated by taking out beads from the reaction mixture, and the rate of bioconversion was assayed. The agar discs were washed with buffer and sterile distilled water after every reaction cycle and reused (Kamble et al, 2013).…”

“…After the initial 10 min, at each 10 min interval, 10 ml of the substrate was added into the column. The reaction was carried out for 110 min when the flow rate was fixed for 1 ml min −1 , 60 min and 45 min when the flow rate was fixed for 2 and 3 ml min −1 , respectively (Kamble et al, 2013).…”

De Novo Genome Assembly of a Plant-Associated Rhodococcus qingshengii Strain (RL1) Isolated from Eruca sativa Mill. and Showing Plant Growth-Promoting Properties

“…Some of these bacteria have large genomes (>5 Mb) with high G+C content (1–4). Their ability to degrade a large spectrum of environmentally problematic compounds (2, 5) or perform quorum quenching (4) makes them suitable for bioremediation or agricultural applications.…”

Nitrile-Degrading Bacteria Isolated from Compost

“…Similarity searches for putative genes involved in nitrile-degradation revealed two genes (BKP42_65670 and BKP42_65680), coding for one of the two putative NHase subunits. These enzymes are known to be responsible for nitrile degradation in various R. erythropolis strains (Kaufmann et al, 1999;Brandão et al, 2003;Vejvoda et al, 2007;Kamble et al, 2013). The deduced protein sequences showed highest identity to the alpha subunit of Rhodococcus sp.…”