FIG. 3.

Particle formation by VP40 and its mutant proteins. The culture medium was harvested 48 h posttransfection with pCEboZVP40, pCEboZVP40AAXY, pCEboZVP40M14A, or pCEboZVP40/1–276, layered over a 20% sucrose cushion, and ultracentrifuged. The pellet resuspended in STE buffer was fractionated through a 10 to 50% discontinuous sucrose gradient. Fractions were collected from the top of the gradient. Fraction proteins were precipitated with 10% TCA and resuspended in SDS-PAGE sample buffer. Total protein from the cell lysate and cleared supernatant (T) was precipitated with 10% TCA and resuspended in SDS-PAGE sample buffer. Proteins were separated by SDS–12% PAGE and detected by Western blotting. Fractions are numbered from the top to the bottom of the gradient.