The catalytic mechanism of amidase also involves nitrile hydrolysis.
The amidase from Rhodococcus rhodochrous J1, which hydrolyzes an amide to an acid and ammonium, was surprisingly found to catalyze the hydrolytic cleavage of the C-N triple bond in a nitrile to form an acid and ammonium stoichiometrically. The amidase exhibited a Km of 3.26 mM for benzonitrile in contrast to that of 0.15 mM for benzamide as the original substrate, but the Vmax for benzonitrile was about 116000 of that for benzamide. A mutant amidase containing alanine instead of Ser195, which is essential for amidase catalytic activity, showed no nitrilase activity, demonstrating that this residue plays a crucial role in the hydrolysis of nitriles as well as amides.
Kobayashi
M
M
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Japan.
Goda
M
M
Shimizu
S
S
D000581:Amidohydrolases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D000641:Ammonia / Q000378:metabolism; D019817:Benzoic Acid / Q000378:metabolism; D002384:Catalysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli; D006868:Hydrolysis; D009570:Nitriles / Q000378:metabolism* / Q000494:pharmacology; D012240:Rhodococcus / Q000201:enzymology*; D013379:Substrate Specificity
D004791:Enzyme Inhibitors; D009570:Nitriles; D000641:Ammonia; D019817:Benzoic Acid; C014356:benzonitrile; D000581:Amidohydrolases; C019665:amidase