Isolation and purification of double-stranded ribonuclease from calf thymus.
A RNase from calf thymus, which specifically cleaves native or synthetic double-stranded RNA molecules endonucleolytically, has been isolated and purified from calf thymus. For optimal activity, the enzyme requires a sulfhydryl reagent and divalent cations; over 95 per cent of the activity is inhibited by 0.5 mm ethidium bromide. The degradation of [3H]poly(C)-poly(I) by purified enzyme preparations yields labeled dinucleotides and octanucleotides; the latter oligonucleotide contained 5'-phosphate and 3'-hydroxyl termini. The enzyme cleaves high molecular weight RNAs such as RNA products formed in vitro by T3 phage-induced RNA polymerase from T3 phage DNA, heterogeneous RNA isolated from duck reticulocyte nuclei, and 45 S RNA isolated from rat liver nucleoli. The mode of degradation of RNA in vitro with the double-stranded RNase is similar to that of Escherichia coli RNase III and appears to act endonucleolytically. The degradation of 45 S RNA with the enzyme results in the production of 29 S and 19 S RNA fragments. These findings suggest that the enzyme may be involved in the processing of high molecular weight precursor RNAs to mRNA or rRNAs in a manner analogous to that reported for RNase III of E. coli.
D000818:Animals; D002417:Cattle; D004355:Drug Stability; D004996:Ethidium / Q000494:pharmacology; D006358:Hot Temperature; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D012260:Ribonucleases / Q000302:isolation & purification / Q000378:metabolism; D013950:Thymus Gland / Q000201:enzymology*
D008345:Manganese; D012260:Ribonucleases; D004996:Ethidium; D008274:Magnesium