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. 2014 Oct 2;6(10):3738–3765. doi: 10.3390/v6103738

Figure 4.

Figure 4

The S40F mutation increases selectively the membrane association of Pr55. (A) HeLa cells were transfected with p∆R PR− HIV-1 constructs directing the expression of either wt Gag, the p6 S40F, S40N, S40D or ∆PTAP mutants or the MA G2A mutant. The cells were lysed by sonication and the post-nuclear supernatant (PNS) was subjected to membrane flotation. Fractions were collected and analyzed by Western blotting for Gag. The Western blot membrane was reprobed with antibodies specific for transferrin receptor (TfR) and β-actin as marker proteins for the membrane or cytosolic fractions, respectively. One representative example of TfR and β-actin blots is shown; (B) Band intensities of the Gag blots were densitometrically quantified and the distribution of Gag was determined for each mutant; (C) The ratio of Gag detected in the membrane fractions (fractions 1 and 2) versus the cytosolic fraction (fractions 4 and 5) was calculated. Values represent the mean of 4 independent experiments ± SD, and were adjusted that wt matches 1.