Purification and biochemical characterization of the carbamate hydrolase from Pseudomonas sp. 50432

A soluble carbamate hydrolase that had a wide specificity was purified 2032-fold from Pseudomonas sp. 50432. This was achieved using a combination of anion-exchange, gel-filtration and hydrophobic-interaction-chromatography techniques. Carbamate hydrolase cleaved the ester linkage of the N-methylcarbamates. The native enzyme was a monomer with a molecular mass of 88 kDa. The optimum pH and temperature of the enzyme activity were 8.5 and 37 °C respectively. The tested cations or EDTA did not affect the enzyme activity. However, 2-mercaptoethanol reversibly inhibited the enzyme activity. The enzyme showed the K m values of 16 and 12 μM for carbofuran and carbaryl respectively. The purified enzyme did not hydrolyse o-nitrophenyl dimethylcarbamate but hydrolysed several N-methylcarbamates and I-naphthyl acetate.