Identification of ubiquitin-like protein-binding subunits of the 26S proteasome.
Ubiquitin-like proteins Rad23 and Dsk2 have recently been shown to be capable of binding both polyubiquitin chains and the 26S proteasome. The ubiquitin-like domains (Ubls) of Rad23 and Dsk2 are indispensable for their interaction with the 26S proteasome, but the proteasome subunits capable of binding the Ubl have not been identified. Here, we report that the Ubls of both Rad23 and Dsk2 can bind with the 19S regulatory particle (RP) of the 26S proteasome in vivo and in vitro. A competition assay using the respective Ubls of Rad23 and Dsk2 revealed that they bind to the RP in a competitive manner. The base subcomplex of the RP was found to have the ability to bind the Ubl. By cross-linking experiments, Rpn1 and Rpn2 were identified as Ubl-binding subunits. Taken together, the results suggest that the Rpn1 and Rpn2 in the base subcomplex form the receptor for the ubiquitin-like protein.
Saeki
Yasushi
Y
Department of Biochemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.
Sone
Takayuki
T
Toh-e
Akio
A
Yokosawa
Hideyoshi
H
D001667:Binding, Competitive; D015153:Blotting, Western; D018797:Cell Cycle Proteins; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D016366:Open Reading Frames; D010447:Peptide Hydrolases / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin / Q000378:metabolism*; D014452:Ubiquitins / Q000737:chemistry / Q000378:metabolism
D018797:Cell Cycle Proteins; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; C100415:DSK2 protein, S cerevisiae; D005656:Fungal Proteins; D011506:Proteins; C490846:RAD23 protein, S cerevisiae; C499663:RPN1 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D014452:Ubiquitins; D005982:Glutathione Transferase; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease