2005
DOI: 10.1128/jvi.79.13.8208-8216.2005
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Recombinant Human Immunodeficiency Virus Type 1 Integrase Exhibits a Capacity for Full-Site Integration In Vitro That Is Comparable to That of Purified Preintegration Complexes from Virus-Infected Cells

Abstract: Retrovirus preintegration complexes (PIC) in virus-Integration of the retrovirus linear DNA genome into host chromosomes is essential for replication of human immunodeficiency virus type 1 (HIV-1). Preintegration complexes (PIC) are formed in the cytoplasm after reverse transcription of the viral RNA. Integrase (IN) in the PIC catalyzes the excision of two nucleotides adjacent to the phylogenetically conserved CA dinucleotide from the 3Ј-OH blunt ends of the viral DNA (7). After nuclear transport of the PIC, t… Show more

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Cited by 75 publications
(94 citation statements)
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“…IN (pNY clone) was expressed in Escherichia coli BL21(DE3) cells and purified to near homogeneity (41). Concentrations of HIV-1 IN were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis using RSV IN as a standard (12), which are very similar to the concentration of HIV-1 IN determined using the molar extinction coefficient of 50,460 M Ϫ1 cm Ϫ1 at 280 nm (24).…”
Section: Methodsmentioning
confidence: 99%
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“…IN (pNY clone) was expressed in Escherichia coli BL21(DE3) cells and purified to near homogeneity (41). Concentrations of HIV-1 IN were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis using RSV IN as a standard (12), which are very similar to the concentration of HIV-1 IN determined using the molar extinction coefficient of 50,460 M Ϫ1 cm Ϫ1 at 280 nm (24).…”
Section: Methodsmentioning
confidence: 99%
“…Integration assay. The integration assay was performed as described previously (41). Briefly, IN was assembled with the donor substrate in presence of 20 mM HEPES (pH 7.0), 5 mM dithiothreitol, 10 mM MgCl 2 , 25 M ZnCl 2 , 100 mM NaCl, and 10% polyethylene glycol (6,000 Da) at 14°C for 15 min.…”
Section: Methodsmentioning
confidence: 99%
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“…2. This essential aspect of the in vivo reaction has typically been difficult to recapitulate in vitro with the purified enzyme, although recent adjustments to assay parameters have boosted efficiency [155,156]. The effect of p75 differed sharply between different recombinant lentiviral IN proteins: its addition enhanced full-site integration for EIAV IN while HIV IN was instead directed virtually exclusively to catalyze uncoupled (half-site) integration in which only one viral DNA end is joined to one strand of target DNA [100].…”
Section: Modeling P75 Function In the Hiv Life Cyclementioning
confidence: 99%
“…Then the full-site (FS) integration occurs when a pair of SC integrates into both strands of target DNA at the opposite sites ( Figure 2A). Besides these, the donor DNA can form other products by binding each other (dimer) or Y-shape Sinha & Grandgenett 2005). To form the SC, different roles were envisioned to describe.…”
Section: Ledgf-ibd Enhanced the Concerted Integration Reaction Of Hivmentioning
confidence: 99%