Comparative biochemical characterization of 5'-phosphodiesterase and phosphomonoesterase from barley malt sprouts.

A comparative biochemical characterization is described of two competing enzymes in the production of flavoring 5'-ribonucleotides, barley malt sprouts 5'-phosphodiesterase (5'-PDE) and phosphomonoesterase (PME). Fractionation of these two enzymes and partial purification of 5'-PDE were achieved by a combination of thermal treatments and precipitation with acetone. With synthetic substrates, under standard assay conditions, 5'-PDE and PME had maximum activities at pH 8.9, 70 degrees C and 55 degrees C, and Km of 0.26 mM and 0.19 mM, respectively. In the presence of 10 mM Mg2+ ions, barley malt sprouts 5'-PDE was activated by up to 160% of the original activity, while PME was inhibited. Zn2+ activated PME by up to 125% of the original activity. Both enzymes were moderately inhibited after addition of Cu2+, Co2+, Ca2+, and Mn2+ ions (10 mM), but, significantly, by addition of the chelating agent EDTA. In the absence of substrate and up to 80 degrees C, barley malt sprouts 5'-PDE showed excellent stability and retained 70% of its original activity at 70 degrees C after 120 min.