Hydration of 3-cyanopyridine to nicotinamide by crude extract nitrile hydratase |
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Authors: | Jacob Eyal Marvin Charles |
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Affiliation: | (1) Bioprocessing Institute, Lehigh University Department of Chemical Engineering, 18015 Bethlehem, PA, USA |
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Abstract: | Summary The kinetic and stability characteristics of crude extract nitrile hydratase fromBrevibacterium R-312 were studied for the hydration of 3-cyanopyridine to nicotinamide. The enzyme was substrate and product inhibited and had the following kinetic constants:Km=28 mM;Kp=36 mM;Ks=155 mM;Vm=5.8 mol/min/mg protein (25°C). Itsmaximum temperature and pH (phosphate buffer) were 35°C and 8.0, respectively and it had half-lives of 50 days, 10 days and 1 day at 4°C, 10°C and 25°C, respectively. The crude extract also exhibited amidase activity on nicotinamide, but it became significant only at nicotinamide concentrations greater than 300 mM. Mathematical models for batch and fed-batch hydrations were developed to account for substrate and product inhibitions and for enzyme decay. They predicted to within 10% experimental results for initial substrate and final product concentrations up to 300 mM; the accuracies decreased at higher concentrations primarily because of the relatively rapid hydrolysis of nicotinamide. |
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Keywords: | Brevibacterium R-312 Nitrile hydratase activity Amidase activity Nitrile hydratase half-life Hydration mathematical models |
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