Proteins are unfolded on the surface of the ATPase ring before transport into the proteasome.
The 19S component of the 26S proteasome contains six ATPase subunits. To clarify how they unfold and translocate proteins into the 20S proteasome for degradation, we studied the homologous archaebacterial proteasome-regulatory ATPase complex PAN and the globular substrate GFP-SsrA. When we attached a small (Biotin) or large (Biotin-Avidin) moiety near its N terminus or a Biotin near its C terminus, GFP-SsrA was unfolded and degraded. However, attaching Avidin near its C terminus blocked passage through PAN and prevented GFP-SsrA degradation. Though not translocated, GFP-Avidin still underwent ATP-dependent unfolding. Moreover, it remained bound to PAN and inhibited further proteolysis. Therefore, (1) translocation and degradation of this substrate require threading through the ATPase in a C to N direction and (2) translocation does not cause but follows ATP-dependent unfolding, which occurs on the surface of the ATPase ring.
Navon
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Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA. ami_navon@hms.harvard.edu
Goldberg
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D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D001105:Archaea / Q000201:enzymology* / Q000378:metabolism; D019843:Archaeal Proteins / Q000737:chemistry* / Q000378:metabolism*; D001360:Avidin / Q000378:metabolism; D001710:Biotin / Q000378:metabolism; D002384:Catalysis; D010447:Peptide Hydrolases / Q000737:chemistry* / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011489:Protein Denaturation; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D019843:Archaeal Proteins; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D001360:Avidin; D001710:Biotin; D000255:Adenosine Triphosphate; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; D000251:Adenosine Triphosphatases