title: Characterization of the adenine nucleoside specific phosphorylase of Bacillus cereus.
abstract: Adenosine phosphorylase, a purine nucleoside phosphorylase endowed with high specificity for adenine nucleosides, was purified 117-fold from vegetative forms of Bacillus cereus. The purification procedure included ammonium sulphate fractionation, pH 4 treatment, ion exchange chromatography on DEAE-Sephacel, gel filtration on Sephacryl S-300 HR and affinity chromatography on N(6)-adenosyl agarose. The enzyme shows a good stability to both temperature and pH. It appears to be a homohexamer of 164+/-5 kDa. Kinetic characterization confirmed the specificity of this phosphorylase for 6-aminopurine nucleosides. Adenosine was the preferred substrate for nucleoside phosphorolysis (k(cat)/K(m) 2.1x10(6) s(-1) M(-1)), followed by 2'-deoxyadenosine (k(cat)/K(m) 4.2x10(5) s(-1) M(-1)). Apparently, the low specificity of adenosine phosphorylase towards 6-oxopurine nucleosides is due to a slow catalytic rate rather than to poor substrate binding.
authors: Dipartimento di Scienze del Farmaco, Università di Sassari, via F. Muroni 23a, 07100 Sassari, Italy. dsfbio@uniss.it
authors: Francesco
authors: F
authors: Sgarrella
authors: Luciano
authors: L
authors: Frassetto
authors: Simone
authors: S
authors: Allegrini
authors: Marcella
authors: M
authors: Camici
authors: Maria Caterina
authors: MC
authors: Carta
authors: Paolo
authors: P
authors: Fadda
authors: Maria Grazia
authors: MG
authors: Tozzi
authors: Piero Luigi
authors: PL
authors: Ipata
mesh_terms: D001409:Bacillus cereus / Q000201:enzymology*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D007700:Kinetics; D008970:Molecular Weight; D011683:Purine-Nucleoside Phosphorylase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: C019603:adenosine phosphorylase; D011683:Purine-Nucleoside Phosphorylase