A novel cold-adapted imidase from fish Oreochromis niloticus that catalyzes hydrolysis of maleimide.
In this paper we report the first comparative study of cold-adapted imidase (EC 3.5.2.2) from the fish (Oreochromis niloticus) liver and its thermophilic counterparts taken from pig liver and Escherichia coli (overexpressed recombinant hydantoinase from Agrobacterium radiobacter NRRL B1). Approximately 6000-fold purification and a 40% yield of fish imidase activity were obtained through ammonium sulfate precipitation, octyl, chelating, DEAE, and hydroxyapatite chromatography. This cold-adapted imidase was characterized by a specific activity 10- to a 100-fold higher than those of its thermophilic counterparts below room temperature (25 degrees C or lower) conditions but less stable at elevated temperatures (40 degrees C or higher). A less organized helical structure (compared to those of pig liver and bacterial imidases) was observed by circular dichroism. Furthermore, maleimide was first identified as a novel substrate of all imidases examined, and confirmed by HPLC and NMR analysis. These results constituted a first study to discover a novel cold-adapted imidase with surprising high activity. These findings might be also helpful for industrial application of imidase.
Huang
Cheng-Yang
CY
Institute of Biochemical Engineering, College of Biological Science and Technology, National Chiao Tung University, Hsinchu, Taiwan, ROC.
Yang
Yuh-Shyong
YS
D000222:Adaptation, Physiological; D000581:Amidohydrolases / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002384:Catalysis; D023681:Cichlids / Q000378:metabolism*; D003080:Cold Temperature; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology / Q000235:genetics; D006868:Hydrolysis; D008099:Liver / Q000737:chemistry / Q000201:enzymology; D008301:Maleimides / Q000737:chemistry*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012231:Rhizobium / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D013379:Substrate Specificity; D013552:Swine; D013696:Temperature
D008301:Maleimides; C043592:maleimide; D000581:Amidohydrolases; C020047:dihydropyrimidinase