Activities of human immunodeficiency virus (HIV) integration protein in vitro: specific cleavage and integration of HIV DNA
Open accessAbstract
Growth of human immunodeficiency virus (HIV) after infection requires the integration of a DNA copy of the viral RNA genome into a chromosome of the host. Here we present a simple in vitro system that carries out the integration reaction and the use of this system to probe the mechanism of integration. The only HIV protein necessary is the integration (IN) protein, which has been overexpressed in insect cells and then partially purified. DNA substrates are supplied as oligonucleotides that match the termini of the linear DNA product of reverse transcription. In the presence of HIV IN protein, oligonucleotide substrates are cleaved to generate the recessed 3' ends that are the precursor for integration, and the cleaved molecules are efficiently inserted into a DNA target. Analysis of reaction products reveals that HIV IN protein joins 3' ends of the viral DNA to 5' ends of cuts made by IN protein in the DNA target. We have also used this assay to characterize the sequences at the ends of the viral DNA involved in integration. The assay provides a simple screen for testing candidate inhibitors of HIV IN protein; some such inhibitors might have useful antiviral activity.
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Sequences internal to the terminal CA and TG dinucleotides are not well conserved, even though they are needed for ideal IN activity during integration.
“However, the sequences internal to these dinucleotides are not well conserved, even though they are needed for ideal IN activity during integration (Bushman and Craigie 1991, Brown 1997, Desfarges and Ciuffi 2010, Ali et al. 2016).”
From Beyond reverse transcription: molecular mechanisms and emerging paradigms in retroviral replication (2026) · cites this paper as Bushman and Craigie 1991
The intasome containing processed viral DNA attaches to host cell DNA, facilitating staggered cleavage of host DNA by IN using viral DNA 3′-hydroxyls as nucleophiles.
“Secondly, the intasome containing the processed viral DNA attaches to the host cell DNA, which facilitates the cleavage of the host cell DNA in a staggered fashion by IN using viral DNA 3′-hydroxyls as nucleophiles (Fujiwara and Mizuuchi 1988, Bushman and Craigie 1991, Engelman et al. 1991).”
From Beyond reverse transcription: molecular mechanisms and emerging paradigms in retroviral replication (2026) · cites this paper as Bushman and Craigie 1991
Cites
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Beyond reverse transcription: molecular mechanisms and emerging paradigms in retroviral replication
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“…Secondly, the intasome containing the processed viral DNA attaches to the host cell DNA, which facilitates the cleavage of the host cell DNA in a staggered fashion by IN using viral DNA 3 -hydroxyls as nucleophiles (Fujiwara and Mizuuchi 1988 , Bushman and Craigie 1991 , Engelman et al. 1991 ).”
- Authors
- Bushman, F. D., & Craigie, R.
- Affiliations
- National Institutes of Health (US)
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 1991-02-15
- Volume
- 88
- Issue
- 4
- Pages
- 1339-1343
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC51013
- OpenAlex
- W2075336365
- Cited by
- 410
- References
- 6
- Citations by year
- 2026: 2 · 2025: 3 · 2024: 2 · 2022: 5 · 2021: 3 · 2020: 7 · 2019: 5 · 2018: 5 · 2017: 5 · 2016: 7 · 2015: 4 · 2014: 6 · 2013: 9 · 2012: 12
- FWCI
- 8.40
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Immunology and Microbiology › Virology › HIV Research and Treatment
- Health Sciences › Medicine › Infectious Diseases › HIV/AIDS drug development and treatment
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Keywords
- HIV-1 integration, viral DNA integration, 3' end cleavage, DNA end joining, HIV integrase inhibitors
- MeSH terms
- Base Sequence, DNA Transposable Elements, DNA Nucleotidyltransferases, DNA, Viral, HIV, Humans, Molecular Sequence Data, Recombinant Proteins, Substrate Specificity, Mutagenesis, Insertional, HIV Long Terminal Repeat, Integrases
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