Figure 2
From: Biochemical and virological analysis of the 18-residue C-terminal tail of HIV-1 integrase
Integrase proteins and in vitro integration assays. (A) Purified proteins (approximately 5 μg each) were stained with Coomassie blue following SDS-polyacrylamide gel electrophoresis. Migration positions of molecular mass standards in kDa are shown on the left. (B) 3' Processing assay. The blunt-ended viral DNA substrate is shown highlighting the subterminal CA that is conserved among all retroviruses, retrotransposons, and some bacterial transposases. During 3' processing, IN cleaves the A/G phosphodiester bond (short vertical arrow), releasing radiolabelled pGTOH dinucleotide. (C) The DNA strand transfer assay utilizes a preprocessed viral DNA end. Integration into target DNA yields products whose lengths exceed that of the starting substrate. (D) Two different DNAs, viral donor (oligonucleotide drawn in the same orientation as in panel C, top) and circular target, are used in the concerted integration assay. In the presence of LEDGFp75, some donor DNA is integrated into only one strand of the target to yield a tagged, nicked circle half-site reaction product. Concerted integration across the major groove by contrast yields a linearized product whose length exceeds that of the starting circle by twice the length of the viral donor. For panels B-D, thin and bold lines represent viral donor and target DNAs, respectively. *, positions of 32P label (panels B and C).