Related Experiment Videos
The sequential mechanism of HIV reverse transcriptase RNase H
M Wisniewski1, M Balakrishnan, C Palaniappan
1Department of Biochemistry and Biophysics and the Cancer Center, University of Rochester, Rochester, New York 14642, USA.
The Journal of Biological Chemistry
|August 25, 2000
Summary
Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase RNase H activity degrades viral RNA. This study reveals that HIV-1 RNase H cleavage sites are independent, with order determined by cleavage rates.
Area of Science:
- Molecular Biology
- Virology
- Enzymology
Background:
- Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase synthesizes viral DNA and degrades the RNA genome via RNase H activity.
- RNase H cleavage of residual RNA fragments occurs in a polymerase-independent manner.
- Previous work established an ordered mechanism for this cleavage, involving primary cuts, secondary/5-nucleotide cuts, and second primary cuts.
Purpose of the Study:
- To investigate the interdependence of cleavage events in the ordered RNase H degradation mechanism of HIV-1.
- To determine if earlier cuts are prerequisites for subsequent cuts in the RNA degradation process.
Main Methods:
- Site-specific inhibition of RNase H cleavage at various positions.
- Analysis of the dependence of later cleavage events on prior cleavage at specific sites.
Main Results:
- Each RNase H cleavage event was found to be independent of other cuts.
- The observed ordered sequence of cleavage is determined by the intrinsic rates of each individual cut, not by sequential dependence.
- A model for unlinked ordered cleavage was proposed.
Conclusions:
- The stepwise degradation of viral RNA by HIV-1 RNase H is an ordered process where individual cleavage events are not dependent on each other.
- The order is dictated by the relative rates of cleavage at different sites.
- This finding provides a mechanistic understanding of viral RNA processing during HIV-1 replication.