Purification and properties of a novel nucleolar exoribonuclease capable of degrading both single-stranded and double-stranded RNA.
A ribonuclease that hydrolyzes either linear duplex or single-stranded RNA in an exonucleolytic manner has been partially purified from Ehrlich ascites tumor cell nucleoli and is free from other ribonucleases. The enzyme will also degrade the RNA complement of an RNA X DNA duplex; however, no nuclease activity is observed on linear duplex or single-stranded DNA. The exonuclease acts on RNA nonprocessively from the 3' end releasing 5'-mononucleotides. The enzyme has a broad pH optimum around pH 8.0, requires Mg2+ or Mn2+ (0.06 mM) for optimum activity, and is sensitive to ethylenediaminetetraacetic acid and N-ethylmaleimide inhibition. Monovalent cations including K+, Na+, and NH4+ are inhibitory. Gel filtration studies of this enzyme gave a Stokes radius of 40 A. Sedimentation velocity measurements in glycerol gradients yield a S20,W of 6.0 S. From these values a native molecular weight of 100 000 was calculated. Copurification of the single- and double-stranded activities, identical reaction requirements, and identical heat-inactivation curves strongly suggest that both activities reside with the same enzyme.
D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000201:enzymology; D002413:Cations, Divalent; D002414:Cations, Monovalent; D002466:Cell Nucleolus / Q000201:enzymology*; D005095:Exoribonucleases / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D011131:Polyribonucleotides; D012330:RNA, Double-Stranded / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D013379:Substrate Specificity
D002413:Cations, Divalent; D002414:Cations, Monovalent; D011131:Polyribonucleotides; D012330:RNA, Double-Stranded; D005095:Exoribonucleases; D012260:Ribonucleases