Integration complexes derived from HIV vectors for rapid assays in vitro
| Autor: | Tal Kafri, Jay S. Siegel, Mark S. T. Hansen, George J. Smith, Valentina Molteni, Frederic D. Bushman |
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| Rok vydání: | 1999 |
| Předmět: |
endocrine system
Anti-HIV Agents Virus Integration Genetic Vectors Biomedical Engineering Integrase inhibitor Bioengineering Applied Microbiology and Biotechnology Polymerase Chain Reaction Fluorescence Viral vector Cell Line Complementary DNA Vector (molecular biology) HIV Integrase Inhibitors DNA Primers biology Base Sequence HIV biochemical phenomena metabolism and nutrition Provirus Virology In vitro Reverse transcriptase Integrase Biochemistry biology.protein Molecular Medicine Biotechnology |
| Zdroj: | Nature biotechnology. 17(6) |
| ISSN: | 1087-0156 |
| Popis: | Of three enzymes encoded by HIV–reverse transcriptase, protease, and integrase—only the first two have been exploited clinically as inhibitor targets. Efforts to develop inhibitors of purified integrase protein have yielded many compounds, but none with clinical utility. A different source of integration activity for studies in vitro is provided by replication intermediates isolated from HIV-infected cells. These preintegration complexes (PICs) can direct integration of the endogenously synthesized viral cDNA into an added target DNA in vitro. Despite their authentic activities, assays of PICs have not been widely used due to technical obstacles, particularly the requirement for handling large amounts of infectious HIV. Here, we describe greatly improved methods for producing PICs using HIV-based vectors that are capable of establishing an integrated provirus but not a spreading infection. We also report the development of a PIC integration assay using DNA-coated microtiter plates, which speeds assays of PIC integration in vitro. We used this method to screen a library of chemicals related to known integrase inhibitors and found a new compound, quinalizarin sulfate, that displayed enhanced activity against PICs. |
| Databáze: | OpenAIRE |
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