Rapid isolation and characterization of the yeast proteasome regulatory complex

The 26S proteasome, which catalyzes degradation of ubiquitinated proteins, is composed of the 20S proteasome and the 19S complex. Recently, it has bee n reported that the 26S complex can be dissociated into the lid complex and the 20S-proteasome-base complex in a mutant yeast and that the lid complex is required for ubiquitin-dependent proteolysis. In the present study, we established methods for rapid isolation of the 19S complex, the lid complex , and the base complex from wild-type yeast. The isolated 19S complex was c apable of binding to the 20S proteasome to reconstitute the 26S proteasome. In contrast with the previously reported result showing that Rpn10, a mult iubiquitin chain binding subunit, is a component of the base complex, we pr esent evidence that the lid complex isolated from wild-type yeast contains Rpn10. (C) 2000 Academic Press.