FIG. 6.

Cosedimentation of inactivated ODC with the 26S proteasome in glycerol density gradient centrifugation. ³⁵S-ODC was incubated for 1 h with a crude extract of HTC cells (640 μg of protein) in the inactivation assay mixture (500 μl), and the mixture was centrifuged at 128,000 × g for 5 h. The precipitates were resuspended and subjected to glycerol density gradient centrifugation as for Fig. 4. Fraction 1 represents the top of the gradient (10% glycerol), and fraction 30 represents the bottom of the gradient (40% glycerol). Samples (0.8 ml) of the gradient fractions were used for determinations of ODC radioactivity, and proteasome activities in the presence and absence of SDS were determined for each 20 μl of the fractions. Arrows indicate the positions of elution of purified 20S and 26S proteasomes.