Isolation and characterization of a cyanide dihydratase from Bacillus pumilus C1.
A cyanide-degrading enzyme from Bacillus pumilus C1 has been purified and characterized. This enzyme consisted of three polypeptides of 45.6, 44.6, and 41.2 kDa; the molecular mass by gel filtration was 417 kDa. Electron microscopy revealed a multimeric, rod-shaped protein approximately 9 by 50 nm. Cyanide was rapidly degraded to formate and ammonia. Enzyme activity was optimal at 37 degrees C and pH 7.8 to 8.0. Activity was enhanced by Sc3+, Cr3+, Fe3+, and Tb3+; enhancement was independent of metal ion concentration at concentrations above 5 microM. Reversible enhancement of enzymatic activity by azide was maximal at 4.5 mM azide and increased with time. No activity was recorded with the cyanide substrate analogs CNO-, SCN-, CH3CN, and N3- and the possible degradation intermediate HCONH2. Kinetic studies indicated a Km of 2.56 +/- 0.48 mM for cyanide and a Vmax of 88.03 +/- 4.67 mmol of cyanide per min/mg/liter. The Km increased approximately twofold in the presence of 10 microM Cr3+ to 5.28 +/- 0.38 mM for cyanide, and the Vmax increased to 197.11 +/- 8.51 mmol of cyanide per min/mg/liter. We propose naming this enzyme cyanide dihydratase.
Meyers
P R
PR
Department of Microbiology, University of Cape Town, South Africa.
Rawlings
D E
DE
Woods
D R
DR
Lindsey
G G
GG
D001407:Bacillus / Q000201:enzymology*; D002413:Cations, Divalent / Q000494:pharmacology; D002499:Centrifugation, Density Gradient; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006867:Hydrolases / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D007700:Kinetics; D046911:Macromolecular Substances; D008345:Manganese / Q000494:pharmacology; D008854:Microscopy, Electron; D008970:Molecular Weight; D011190:Potassium Cyanide / Q000378:metabolism; D013816:Thermodynamics
D002413:Cations, Divalent; D046911:Macromolecular Substances; D008345:Manganese; D006867:Hydrolases; C083488:cyanide dihydratase; D011190:Potassium Cyanide