Yeast total tRNA inhibits Gag binding to PC + PS liposomes. [35S]-labeled Gag synthesized using rabbit reticulocyte lysates was treated (or left untreated) with RNase A at 37°C for 20 min. After inactivation of RNase A with RNasin, the mixtures were further incubated with varying concentrations of yeast tRNA at 37°C for 30 min. Subsequently, PC + PS liposomes were added and incubated further for 15 min before performing equilibrium flotation centrifugation. Fractions were analyzed using SDS-PAGE followed by phosphorimager analysis. (M) Membrane-bound, (NM) non-membrane-bound. The liposome binding efficiency was calculated as the amount of membrane-bound Gag as a fraction of total Gag. Data from three different experiments are shown as mean ± standard deviation. P-values were determined in comparison to the no RNA add-back condition using Student's t-test. (**) P < 0.01; (***) P < 0.001. Reduction of liposome binding efficiency to 50% was observed when RNase-treated Gag was incubated with 91 ng/µL yeast tRNA prior to mixing with liposomes.
