Purification, stability and kinetic properties of highly purified adenosine deaminase from Bacillus cereus NCIB 8122

• Published in: Biochimica et biophysica acta Vol. 884; no. 3; pp. 490 - 496
• Main Authors: Gabellieri, Edi, Bernini, Simonetta, Piras, Luciana, Cioni, Patrizia, Balestreri, Ettore, Cercignani, Giovanni, Felicioli, Romano
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 10.12.1986; Elsevier; North-Holland
• Subjects: Adenosine deaminase; Adenosine Deaminase - isolation & purification; Adenosine Deaminase - metabolism; Analytical, structural and metabolic biochemistry; B. cereus; Bacillus cereus - enzymology; Biological and medical sciences; Cations, Monovalent; Enzyme Stability; Enzymes and enzyme inhibitors; Fundamental and applied biological sciences. Psychology; Glycerol; Glycerol - pharmacology; Hydrolases; Kinetics; Monovalent cation; Nucleoside Deaminases - isolation & purification; Glycerol; B. cereus; Monovalent cation; Adenosine deaminase; Bacillus cereus; Purification; Bacillales; Gel electrophoresis; Enzyme; Bacteria; Bacillaceae; Catalyst activity; Physicochemical properties
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/0304-4165(86)90199-6

Adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) from Bacillus cereus NCIB 8122 has been purified to electrophoretic homogeneity by ammonium sulfate precipitation, gel filtration through Sephadex G-100, DEAE-Sephadex A-50 chromatography and ion-exchange HPLC on DEAE-Polyol. The enzyme acitivity is stabilized (at temperatures from 0°C to 40°C) by 50 mM NH 4 + of K +, while it is irreversibly lost in the absence of these or a few other monovalent cations. Glycerol (24% by volume) helps tha cation in stabilizing the enzyme activity above 40°C, but also exerts per se a noticeable protecting effect at room temperature. B. cereus adenosine deaminase displays the following properties: M r on Sephadex G-200, 68 000; M r in SDS-polyacrylamide gel electrophoresis, 53 700; optimal pH-stability (in the presence of 50 mM KCl) over the range 8–11 at 4°C, and maximal catalytic activity at 30°C between pH 7 and 10; K m for adenosine around 50 μM over the same pH range and K m for 2′-deoxyadenosine around 400 μM.