ArticleNucleic acids research2002
Assembly, purification and crystallization of an active HIV-1 reverse transcriptase initiation complex.
Article in Nucleic acids research, 2002. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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Who cites it
4 citing papers in PubMed, 6 citations in OpenAlex.
- General Strategies for RNA X-ray Crystallography.Molecules (Basel, Switzerland) · 2023Review
- Article
- Structure of HIV-1 reverse transcriptase bound to an inhibitor active against mutant reverse transcriptases resistant to other nonnucleoside inhibitors.Proceedings of the National Academy of Sciences of the United States of America · 2004Article
- Synthetic tRNALys,3 as the replication primer for the HIV-1HXB2 and HIV-1Mal genomes.Nucleic acids research · 2004Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) initiates DNA synthesis from the 3' end of human tRNA(Lys3). We have used cis-acting hammerhead ribozymes to produce homogeneous-length transcribed tRNA(Lys3) and have developed conditions for purifying highly structured RNAs on a modified tube-gel apparatus. Titration experiments show that this RNA can assemble into an initiation complex that contains equimolar amounts of HIV-1 RT, transcribed tRNA(Lys3), and chemically synthesized template RNA. We have purified this complex using gel-filtration chromatography and have found that it is homogeneous with respect to molecular weight, demonstrating that the initiation complex forms a single discrete species at micromolar concentrations. When this initiation complex is supplied with deoxynucleotides, essentially all of the tRNA is used as a primer by HIV-1 RT and is fully extended to the 5' end of the template. Thus, in vitro transcribed tRNA can be used efficiently as a primer by HIV-1 RT. We have also obtained crystals of the HIV-1 initiation complex that require the precisely defined ends of this in vitro transcribed tRNA(Lys3) to grow.
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