Purification and properties of human urine ribonucleases

1. Two RNases (RNase UL and RNase US) were purified from the urine of human adults by means of column chromatographies on SP-Sephadex C-50, phospho-cellulose and CM-cellulose and gel-filtration on Sephadex G-75 in homogeneous states obtained by SDS-disc electrophoresis. 2. Molecular weights of these RNases determined by gel-filtration were 38,000 and 13,000 for RNase UL and RNase US, respectively. 3. Optimal pH's of urine RNases were 8.0 and 6.75 for RNase UL and RNase US, respectively. 4. Chemical composition of urine RNases was determined. RNase UL contains about 20.7% of neutral sugar and 7.8% of hexosamine. RNase US contains a very small amount of carbohydrate moiety. 5. Base specificity of urine RNases studied with 2',3'-cyclic nucleotides and dinucleoside phosphates as substrates indicated that both RNases were pyrimidine specific and cytosine preferential enzyme, as is bovine pancreatic RNase A. Although base specificity of RNase UL was qualitatively similar to RNase A, that of RNase US was slightly different. That is, RNase US did not hydrolyze UpU and hydrolyzed UpC and 2',3'-cyclic UMP very slowly. 6. Antigenic properties of human urine RNases were studied by Ouchterlony's double diffusion analysis. RNase UL, RNase US, and RNase A were serologically distinguishable.