SELF-CLEAVAGE OF HEPATITIS DELTA-VIRUS GENOMIC STRAND RNA IS ENHANCED UNDER PARTIALLY DENATURING CONDITIONS

Self-cleavage of a polyribonucleotide containing an autocleaving sequence from the genomic strand of hepatitis delta virus was enhanced by conditions that destabilized RNA structure. Self-cleavage of the transcripts used in this study required Mg2+ (or another divalent cation), and in the absence of denaturants, maximum cleavage was observed at very low Mg2+ concentrations (0.05–0.1 mM). However, at 37 °C and in the presence of 2–10 mM Mg2+ the rate of cleavage was increased as much as 50-fold with the addition of urea to 5 M or formamide to 10 M. Cleavage was prevented by higher concentrations of the same reagents (9.5 M urea or 22.5 M formamide), presumably because a structure required for selfcleavage is disrupted by strongly denaturing conditions. In contrast to a previous report [Wu, H.-N., & Lai, M. M. C. (1989) Science 243, 652–654], we find that chelating Mg2+ with EDTA terminates the cleavage reaction without promoting measurable amounts of ligation of the cleavage products. The ability of denaturants to promote rapid self-cleavage in vitro raises the possibility that an unidentified factor could have a similar effect in vivo. © 1990, American Chemical Society. All rights reserved.