title: A novel L-asparaginase from Aquabacterium sp. A7-Y with self-cleavage activation.
abstract: We have identified a novel L-asparaginase, abASNase3, from Aquabacterium sp. A7-Y. abASNase3 is composed of 306 amino acids and exhibits 34 % sequence homology to human asparaginase (hASNase3). Further analysis revealed that abASNase3 belongs to the N-terminal nucleophile (Ntn) family of hydrolases. Previous reports about the Ntn hydrolase family and the results of our study suggest that abASNase3 must form two subunits by self-cleavage between Gly189 and Thr190 to attain catalytic activity. The two subunits remained tightly associated to build a single functional unit. The optimum pH for abASNase3 was found to be 8.0 in Tris-HCl buffer and the enzyme was found to be stable over a broad pH range from pH 6.0 to 12.0. The optimum temperature for abASNase3 was found to be approximately 40 °C, and the enzyme was stable below 65 °C. abASNase3 showed high substrate specificity toward L-asparagine and had no or only slight activity toward D-asparagine, L-glutamine and D-glutamine. abASNase3 was significantly activated by Mg(2+) and was substantially inhibited by Ni(2+), Cu(2+), Mn(2+) and Co(2+). The Michaelis-Menten constant and turnover number of abASNase3 for L-asparagine were estimated to be 3.37 × 10(-2) M and 8.72 × 10(-3) s(-1), respectively. Our results indicate that abASNase3 is a novel L-asparaginase in the Ntn family of hydrolases.
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China. zhibin.sun@yahoo.com.
authors.forename: Zhibin
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors.forename: Ding
authors.initials: D
authors.lastname: Li
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors.forename: Pingping
authors.initials: P
authors.lastname: Liu
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors.forename: Wenhui
authors.initials: W
authors.lastname: Wang
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors.forename: Kai
authors.initials: K
authors.lastname: Ji
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Key Laboratory of Environmental Microbiology of Ministry of Agriculture, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China. czl@njau.edu.cn.
authors.forename: Zhongli
authors.initials: Z
authors.lastname: Cui
mesh_terms: D000595:Amino Acid Sequence; D001215:Asparaginase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D019117:Burkholderia / Q000201:enzymology*; D002850:Chromatography, Gel; D057075:Enzyme Assays; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017385:Sequence Homology; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D008670:Metals; D011994:Recombinant Proteins; D005973:Glutamine; D001215:Asparaginase
keywords: Aquabacterium sp. A7-Y; L-Asparaginase; Ntn family of hydrolases; Self-cleavage activation