title: Overproduction, Purification and Characterization of Adenylate Deaminase from Aspergillus oryzae.
abstract: Adenylate deaminase (AMPD, EC 3.5.4.6) is an aminohydrolase that widely used in the food and medicine industries. In this study, the gene encoding Aspergillus oryzae AMPD was cloned and expressed in Escherichia coli. Induction with 0.75 mM isopropyl β-D-l-thiogalactopyranoside resulted in an enzyme activity of 1773.9 U/mL. Recombinant AMPD was purified to electrophoretic homogeneity using nickel affinity chromatography, and its molecular weight was calculated as 78.6 kDa. Purified AMPD exhibited maximal activity at 35 °C, pH 6.0 and 30 mM K+, with apparent Kmand Vmaxvalues of 2.7 × 10-4M and 77.5 μmol/mg/min under these conditions. HPLC revealed that recombinant AMPD could effectively catalyse the synthesis of inosine-5'-monophosphate (IMP) with minimal by-products, indicating high specificity and suggesting that it could prove useful for IMP production.
authors.affiliation: College of Light Industry and Food Engineering, Guangxi University, Nanning, 530004, China.
authors.forename: Shubo
authors.initials: S
authors.lastname: Li
authors.affiliation: College of Life Science and Technology, Guangxi University, Nanning, 530004, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Qian
authors.affiliation: College of Life Science and Technology, Guangxi University, Nanning, 530004, China.
authors.forename: Yunlong
authors.initials: Y
authors.lastname: Liang
authors.affiliation: College of Life Science and Technology, Guangxi University, Nanning, 530004, China.
authors.forename: Xinkuan
authors.initials: X
authors.lastname: Chen
authors.affiliation: College of Light Industry and Food Engineering, Guangxi University, Nanning, 530004, China.
authors.forename: Mouming
authors.initials: M
authors.lastname: Zhao
authors.affiliation: National Engineering Research Center for Non-Food Biorefinery, Guangxi Academy of Sciences, Nanning, 530004, China. guoyuan@gxas.cn.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Guo
authors.affiliation: College of Life Science and Technology, Guangxi University, Nanning, 530004, China. pangzw@gxu.edu.cn.|College of Life Science and Technology, Guangxi University, Nanning, 530005, China. pangzw@gxu.edu.cn.
authors.forename: Zongwen
authors.initials: Z
authors.lastname: Pang
mesh_terms: D000659:AMP Deaminase / Q000302:isolation & purification* / Q000378:metabolism*; D000249:Adenosine Monophosphate / Q000737:chemistry / Q000378:metabolism; D001236:Aspergillus oryzae / Q000187:drug effects / Q000201:enzymology*; D055162:Biocatalysis / Q000187:drug effects; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007291:Inosine Monophosphate / Q000737:chemistry / Q000378:metabolism; D007477:Ions; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000378:metabolism; D013696:Temperature
chemical_list: D007477:Ions; D008670:Metals; D011994:Recombinant Proteins; D007291:Inosine Monophosphate; D000249:Adenosine Monophosphate; D000659:AMP Deaminase
keywords: Adenylate deaminase; Aspergillus oryzae; Catalytic properties; Inosine-5ʹ-monophosphate; Purification