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 degrees 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 self-cleavage 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.
Rosenstein
S P
SP
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710.
Been
M D
MD
D001483:Base Sequence; D005559:Formamides / Q000494:pharmacology; D003698:Hepatitis Delta Virus / Q000235:genetics*; D008274:Magnesium / Q000502:physiology; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D012323:RNA Processing, Post-Transcriptional / Q000187:drug effects; D016337:RNA, Catalytic / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics*; D014508:Urea / Q000494:pharmacology
D005559:Formamides; D016337:RNA, Catalytic; D012330:RNA, Double-Stranded; D012367:RNA, Viral; C031066:formamide; D014508:Urea; D008274:Magnesium