Purification and Properties of Porcine Pancreatic Ribonuclease

Abstract

1. A ribonuclease (RNase P) was isolated from porcine pancreas. The enzyme was purified by means of ammonium sulfate fractionation, phenol extraction, DEAE-, Phospho- and CM-cellulose column chromatography about 850 fold. 2. Purified RNase P was homogeneous in centrifugation (S=2.10). The specific activity of RNase P was about 70–85% of RNase A [EC 2.7.7.16, ribonucleate pyrimidinenucleotido-2'-transferase (cyclizing)] under the conditions used for estimation. 3. RNase P was very similar to that of RNase A in respect to molecular weight, heat stability and the activity dependence on ionic strength. 4. The substrate specificity of the enzyme was qualitatively the same as that of RNase A and the enzyme was inhibited by Cu⁺⁺, Zn⁺⁺ and Hg⁺⁺ as in the case of RNase A. 5. The pH optima of RNase P were at pH 7.5 (RNA as substrate) and 6.0 (cytidine-2', 3'-cyclic phosphate (C-cyclic-P) as substrate). 6. The isoelectric point and elution positions from the columns (CM- and Phospho-cellulose) of RNase P indicated that this ribonuclease was less basic than RNase A.