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Characterization of the double stranded RNA dependent RNase activity associated with recombinant reverse
H Ben-Artzi1, E Zeelon, S F Le-Grice
1BioTechnology General Ltd, Kiryat Weizmann, Rehovot, Israel.
Nucleic Acids Research
|October 11, 1992
Summary
This study reveals that human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) possesses RNA/RNA dependent RNase activity. This activity, distinct from RNaseH, suggests overlapping catalytic sites for RNA degradation.
Area of Science:
- Molecular Biology
- Virology
- Enzymology
Background:
- Reverse transcriptase (RT) is crucial for retroviral replication.
- RTs possess RNaseH activity, essential for viral DNA synthesis.
- The presence of other RNase activities in RTs is less understood.
Purpose of the Study:
- To investigate RNA/RNA dependent RNase activity in HIV-1 RT.
- To characterize the enzyme's active sites involved in RNA degradation.
- To differentiate RNA/RNA dependent RNase activity from RNaseH activity.
Main Methods:
- An in situ gel assay using [32P] RNA/RNA substrate was developed.
- Proteins were separated by electrophoresis, renatured, and enzymatic activity was assessed.
- Purified HIV-1 RT and mutant forms were analyzed.
Main Results:
- HIV-1 RT p66/p51 exhibited RNA/RNA dependent RNase activity, distinct from RNaseH.
- This activity comigrated with the p66 subunit and was mapped to the carboxy-terminal region.
- A contaminating E. coli RNaseIII-like enzyme was also observed.
Conclusions:
- HIV-1 RT possesses intrinsic RNA/RNA dependent RNase activity.
- The findings suggest overlapping or identical active sites for RNA/DNA and RNA/RNA degradation.
- This activity may play a role in viral replication or pathogenesis.