RNase footprinting demonstrates antigenomic hepatitis delta virus ribozyme structural rearrangement as a result of self-cleavage reaction.

BMC Research Notes
L P SavochkinaR Sh Beabealashvilli

Abstract

Hepatitis delta virus (HDV) is a satellite virus of hepatitis B. During viral replication the 1700-nucleotide-long genomic RNA and its complement, the antigenomic RNA, undergo self-cleavage catalyzed by internal ribozyme motifs that are essential for propagation of the virus in vivo. These self-cleavage activities are provided by 85-nucleotide-long sequence elements, the genomic and antigenomic forms of HDV ribozyme. Recently four permuted variants of the antigenomic HDV cis-ribozyme with a self-cleavage site located at the 5' proximity, in the middle, or nearby the 3' end of the molecule were constructed and synthesized. These constructs exhibit equal activity, a bi-phasic kinetics of self-cleavage reaction and reaction products with low and high stability. We have used ribonuclease probing to footprint the structures of uncleaved and post-cleaved forms of the antigenomic HDV ribozymes in solution. Uncleaved ribozymes, associated and individual products of the self-cleavage reaction were analyzed using ribonuclease and Fe(II)-EDTA protection assays to reveal the differences in the structure of pre- and post-cleaved antigenomic HDV ribozyme in solution. Our findings demonstrate that a significant conformational change accompani...Continue Reading

References

Aug 1, 1997·European Journal of Biochemistry·M D Been, G S Wickham
Oct 23, 1998·Nature·A R Ferré-D'AmaréJ A Doudna
Dec 11, 1999·Nucleic Acids Research·H M BermanP E Bourne
Dec 4, 2003·Biochemistry. Biokhimii︠a︡·T I BeliankoR Sh Beabealashvilli

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Citations

Nov 10, 2009·Nature Structural & Molecular Biology·Nadia KulshinaAdrian R Ferré-D'Amaré
Jan 31, 2012·RNA·Hari BhaskaranJohn J Perona

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Methods Mentioned

BETA
footprinting
electrophoresis
X-ray

Software Mentioned

RasMol
Tripos Sybyl
RasMol script

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