On RNase and other hydrolytic enzymes in excised Avena leaf tissues

As compared to the zero-time control, 40, 80 and 170% increases in phospho-monoesterase, phosphodiesterase and ribonuclease (RNase) activities were found in excised Avena leaf tissues incubated in the light in a wet chamber for 7.5 hr. The increase in enzyme activity was completely inhibited by cycloheximide. A ribonuclease, an acidic and an alkaline phosphodiesterase were partially purified from control and excised Avena leaf tissues. The RNase, purified about 150 fold, was characterized in detail. It is localized in the soluble fraction. The enzyme has a pH-optimum of 5.5 with yeast RNA as substrate. It is heat sensitive. Bivalent cations and p-chloromercuribenzoate are potent inhibitors of enzyme activity. The enzyme hydrolyzes poly A, poly C, poly U and sRNA. The reaction rate is the highest with poly A as substrate. Native or heat-denatured DNA, double stranded poly A-poly U hybrid and bis-pnitrophenylphosphate are not attacked. The RNase hydrolyzes purine 2′,3′-cycIic phosphates. The pyrimidine 2′,3′-cyclic phosphates are not attacked. Enzyme activity is subjected to product inhibition. 2′(3′)-AMP, 2′(3′)-GMP, adenosine 2′,3′-cyclic phosphate and guanosine 2′,3′-cyclic phosphate are potent inhibitors. 2′(3′)-CMP, 2′(3′)-UMP, cytidine 2′,3′-cyclic phosphate and uridine 2′,3′-cyclic phosphate have negligible or no inhibitory effects. On the basis of these properties the enzyme is classified as an RNase exhibiting a relative purine specificity. The above properties of RNases isolated from intact and excised leaves proved to be identical.