Figure 1
From: Re-visiting the functional Relevance of the highly conserved Serine 40 Residue within HIV-1 p6Gag
HIV-1 p6 Gagvariants and their effect on Gag processing and viral release. (A) Scheme of the gag and pol ORFs in the HIV-1NL4-3unc genome [15]. The arrow with an asterisk indicates the frameshift signal at the 3’ end of gag. Amino acid sequences of the altered p6 sequences of the NL4-3unc variants are shown below; mutated residues are indicated. The NL4-3 late(-) and Vpr(-) variants were constructed in the wild-type pNL4-3 proviral plasmid with overlapping gag and pol reading frames. (B) Gag processing and particle release efficiency. Virus particles were prepared by ultracentrifugation from the culture media of HEK 293 T cells transfected with the indicated proviral plasmids. Cell lysates (top panel) and virus particles (bottom panel) were analyzed for Gag-derived products by quantitative immunoblot (LiCor) using antiserum against HIV-1 CA. Positions of molecular mass standards are shown to the left, Gag-derived processing products are indicated to the right. (C) Quantitative immunoblotting of cell lysates and particles as shown in (B) was used to calculate the amount of released CA-containing proteins by quantification of band intensities using the LiCor Odyssey 3.0 software. Relative release was calculated by dividing the sum of CA-reactive band intensities in the particle fraction by total CA-reactive band intensities in cell lysates and particles. Mean values and SD from three independent experiments performed in duplicates are shown. p-values were calculated using an unpaired student’s t-test (n.s. > 0.05, * < 0.05, *** < 0.001).