The roles in the retroviral replication are firmly established and have been examined in increasingly great detail. Moreover, RNase H constitutes an essential functional domain of the reverse transcriptases of retroviruses, hepadnaviruses (HBV RT) and transposons ( 3). This activity was first observed in calf thymus extracts ( 1) then recognized in all organisms examined to date in all three kingdoms ( 2). Ribonucleases H cleave specifically the RNA strand of RNA–DNA hybrids. Finally, the inhibitory aptamers were capable of completely abolishing the action of an antisense oligonucleotide in a rabbit reticulocyte lysate supplemented with human RNase H1, with IC 50 ranging from 50 to 100 nM. Base pairing between the 5′ and 3′ tails appears crucial for conferring the inhibitory properties to the aptamer. The VI‐2 structure consists of a central region unimolecular quadruplex formed by stacking of two guanine quartets flanked by the 5′ and 3′ tails that form a stem of six base pairs.
The two inhibitory oligomers, V‐2 and VI‐2, were quite different in structure with V‐2 folding into a large, imperfect but stable hairpin loop. Only two of the 37 DNA aptamers selected were inhibitors of human RNase H1 activity. None of them could fold into a simple perfect double‐stranded DNA hairpin confirming that double‐stranded DNA does not constitute a trivial ligand for the enzyme. Using SELEX, we have generated a set of DNA sequences that can bind efficiently ( K d values ranging from 10 to 80 nM) to the human RNase H1.
Human RNase H1 binds double‐stranded RNA via its N‐terminal domain and RNA–DNA hybrid via its C‐terminal RNase H domain, the latter being closely related to Escherichia coli RNase HI.
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