17] Double-stranded RNA nuclease associated with rye germ ribosomes

Abstract

Publisher Summary This chapter describes a two-step procedure to isolate and purify a novel nuclease able to degrade double-stranded RNA, and some properties of this enzyme (action on single- and double-stranded RNAs, optimum pH, effects of ionic strength and cations on enzyme activity, and hydrolysis of polynucleotides). Data on the ability of the nuclease to cleave Lupinus luteus 5S rRNA is also presented. The enzyme is called rye germ ribosomal nuclease II, to distinguish it from nuclease I, the enzyme previously purified from rye germ ribosomal washes. The two methods yielded similar results: the nuclease preparations showed a specific activity toward poly(I) · poly(C) about 30-fold higher than the nuclease present in the ribosomes. In addition to its activity toward dsRNA and ssRNA, nuclease II cleaves native and denatured DNA. These results indicate that it is a sugar-nonspecific nuclease. Like other plant nucleases rye germ nuclease II is most active toward poly(U) and poly(A). Nuclease I, previously isolated from an ammonium chloride wash of rye germ ribosomes, 4 is most active toward poly(C). It is conceivable that nuclease II associated with rye germ ribosomes may be involved in posttranscriptional silencing.