Purification and some properties of cytosol 5'-nucleotidase from rat liver.

A 5'-nucleotidase (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) was highly purified from rat liver. The preparation appeared homogeneous on the criteria of disc-gel electrophoresis. A pH optimum at about 6.5 was observed for all substrates tested. The activity of this enzyme was absolutely dependent on the presence of various bivalent metal salts. The highest V value was attained with MgCl2 and the concentration at half-enzyme saturation was lowest with MnCl2. The enzyme had markedly higher affinities for IMP, dIMP, GMP and dGMP than the other 5'-mononucleotides, although V values for all the substrates tested were in the same order of magnitude. The activity of this enzyme was stimulated by various alkali metal salts, some carboxylic acids and adenine nucleotides. When AMP was used as substrate, the substrate-velocity plot was sigmoidal and NaCl, Tris-maleate and ATP stimulated the enzyme by decreasing the sigmoidicity of the plot. When IMP was used as substrate, the substrate-velocity plot was hyperbolic and these three activators stimulated the enzyme by increasing the V and decreasing the Km value. Some of these results provided consistent evidence for the identity of this enzyme and the cytosol 5'-nucleotidase, the presence of which had been reported in crude preparations from rat liver.