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Figure 2 | Retrovirology

Figure 2

From: HuR interacts with human immunodeficiency virus type 1 reverse transcriptase, and modulates reverse transcription in infected cells

Figure 2

The HIV-1 RT heterodimer interacts directly with GST-HuR. A. Coomassie blue staining of the purified RT heterodimer. NP: non purified, NF: non fixed, E1–E3: elutions. B. Coomassie blue staining of the purified GST and GST-HuR. NP: non purified, P: purified. C. Schematic representation of our HTRF assay (adapted from Cisbio Intl.). Europium trisbipyridine cryptate (TBPEu3+) was coupled to anti-GST antibodies, acting as the FRET energy donor, following excitation at 337 nm. Cross-linked allophycocyanin (XL665) was coupled to anti-His antibodies acting as the FRET energy acceptor and emitting a sustained signal at 665 nm. D. Serial dilutions of purified GST-HuR or GST alone were incubated with a constant concentration of RT-His (10 ng/ml), in the presence of constant amounts of anti-His-XL665 and anti-GST-TBPEu3+ antibodies. The Fret signal was measured after 24 hours of incubation at 4°C. These results are representative of those obtained in four independent experiments.

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