Propertles of 5'-nucleotidase from Bacillus Cereus obtained by washing intact cells with water

5'-Nucleotidase activity can be detected in intact cells of Bacillus cereus and is released simply by washing with aqueous media; the release is partly dependent upon storage of cells at —20°, but is scarcely affected by the solute composition of the washing medium. The enzyme preparation obtained by washing cells catalyzes the dephosphorylation of various 5’¬mononucleotides: IMP (Km = 40 μM) is hydrolyzed 5 times faster than AMP (Km = 4 μM); dAMP (Km = 7.5 μM) is also dephosphorylated at a higher rate than AMP. The pyrophos¬phate linkage of ADP-glucose and NAD+ is also hydrolyzed. ADP and ATP, which are noi attacked by B. cereus 5'-nucleo¬tidase, exert a strong inhibition on AMP and dAMP hydrolysis (Ki = 2 and 5 μM respectively): other nucleoside triphosphates are also inhibitory. AMP hydrolysis catalyzed by intact cells of B. subtilis is also sensitive to ATP and ADP inhibition. The optimal pH for adenine nucleotide hydrolysis is around 8.4; divalent cations, such as Ca2+, Mg2+, Mn2+, and CO2+, activate 5'-nucleotidase below pH 8, thus shifting the optimal pH to less alkaline values. Activation by Mg2+ and Ca2+ at pH 7.2 is abolished in the presence of ATP, whose inhibitory effect is strengthened by the divalent cation; on the other hand, ADP inhibition and divalent cation activation behave as independent effects at pH 7.2. The apparent surface location of the enzyme and its kinetic properties are interpreted as indicating its potential role in the uptake of extracellular nucleic acid material by chemical conversion into compounds which can penetrate the cell membrane, without affecting the cytoplasmic nucleotide pool.