FIG. 4.
Kinetic analysis of wt and mutant Gag processing. (a) Kinetics of wt Gag processing and virus release. HeLaP4 cells were transfected with pMPMV and pulse-labeled with [35S]methionine for 30 min. Cells were chased in DMEM for the indicated time. Cell- and virus-associated CA-containing proteins (lanes 1 to 6 and 7 to 12, respectively) were immunoprecipitated with anti-CA antiserum, resolved by SDS-PAGE, and visualized by phosphorimage analysis. (b) wt and L-domain mutant processing kinetics. HeLaP4 cells were transfected with the indicated pMPMV constructs, labeled as described above, and chased for 0, 2, or 8 h. At the 2-h time point, a Gag-related band (arrow) that migrates slower than Gag was detected in cells expressing PPPY mutant and double mutant Gag.