Nucleic Acids Research, Vol 25, Issue 11 2167-2173, Copyright © 1997 by Oxford University Press
U Koppelhus, V Zachar, PE Nielsen, X Liu, J Eugen-Olsen and P Ebbesen
We have tested the inhibitory potential of peptide nucleic acid (PNA) on in
vitro reverse transcription of the HIV-1 gag gene. PNA was designed to
target different regions of the HIV-1 gag gene and the effect on reverse
transcription by HIV-1, MMLV and AMV reverse transcriptases (RTs) was
investigated. We found that a bis-PNA (parallel antisense 10mer linked to
antiparallel antisense 10mer) was superior to both the parallel antisense
10mer and antiparallel antisense 10mer in inhibiting reverse transcription
of the gene, thus indicating triplex formation at the target sequence. A
complete arrest of reverse transcription was obtained at approximately
6-fold molar excess of the bis-PNA with respect to the gag RNA. At this
molar ratio we found no effect on in vitro translation of gag RNA. A 15mer
duplex- forming PNA was also found to inhibit reverse transcription at very
low molar ratios of PNA/ gag RNA. Specificity of the inhibition of reverse
transcription by PNA was confirmed by RNA sequencing, which revealed that
all tested RTs were stopped by the PNA/RNA complex at the predicted site.
We propose that the effect of PNA is exclusively due to steric hindrance,
as we found no signs of RNA degradation that would indicate PNA-mediated
RNase H activation of the tested RTs. In conclusion, PNA appears to have a
potential to become a specific and efficient inhibitor of reverse
transcription in vivo , provided sufficient intracellular levels are
achievable.
ARTICLES
Efficient in vitro inhibition of HIV-1 gag reverse transcription by peptide nucleic acid (PNA) at minimal ratios of PNA/RNA
Department of Virus and Cancer, Danish Cancer Society, Gustav Wieds Vej 10, DK-8000 Aarhus C, Denmark. ukop@biobase.dk
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