Purification and characterization of a ribonuclease from human spleen. Immunological and enzymological comparison with nonsecretory ribonuclease from human urine.
A ribonuclease has been isolated from human spleen (RNase HS) by means of acid extraction, ammonium sulphate fractionation, successive column chromatographies on CM-cellulose, heparin-actigel, and poly(G)-agarose, and double gel-filtration on Sephadex G-75. The purified preparation was homogeneous as judged by SDS/PAGE. RNase HS was found to be a glycoprotein, containing three fucose, one mannose and five glucosamine residues/molecule, with a molecular mass of 17 kDa as determined by both SDS/PAGE and gel filtration. The catalytic properties and structural features, including its amino acid composition and the amino acid sequence of the N-terminal 35 residues, indicated that the enzyme was strictly related to nonsecretory RNase isolated from human urine and liver. In particular, the amino acid sequence of the N-terminal was identical with that of urine nonsecretory RNase and eosinophil-derived neurotoxin. Furthermore, analyses using three different antibodies specific to RNase HS, urine nonsecretory RNase and urine secretory RNase, indicated that RNase HS was not immunologically distinguishable from urine nonsecretory RNase, but clearly so from urine secretory RNase. However, the carbohydrate compositions of RNase HS and urine nonsecretory RNase were found to differ. It therefore remains to be resolved whether or not the tissue of origin of nonsecretory RNase in urine is the spleen.
Yasuda
T
T
Department of Legal Medicine, Fukui Medical School, Japan.
Mizuta
K
K
Sato
W
W
Kishi
K
K
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000906:Antibodies / Q000276:immunology; D002384:Catalysis; D002845:Chromatography / Q000379:methods; D002850:Chromatography, Gel; D004722:Endoribonucleases / Q000276:immunology / Q000302:isolation & purification* / Q000652:urine; D005260:Female; D006023:Glycoproteins / Q000276:immunology / Q000302:isolation & purification; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D013154:Spleen / Q000201:enzymology*
D000596:Amino Acids; D000906:Antibodies; D006023:Glycoproteins; D004722:Endoribonucleases; C029494:ribonuclease U