Recognition of misfolding proteins by PA700, the regulatory subcomplex of the 26 S proteasome.
The 26 S proteasome is a large protease complex that catalyzes the degradation of both native and misfolded proteins. These proteins are known to interact with PA700, the regulatory subcomplex of the 26 S proteasome, via a covalently attached polyubiquitin chain. Here we provide evidence for an additional ubiquitin-independent mode of substrate recognition by PA700. PA700 prevents the aggregation of three incompletely folded, nonubiquitinated substrates: the DeltaF-508 mutant form of cystic fibrosis transmembrane regulator, nucleotide binding domain 1, insulin B chain, and citrate synthase. This function does not require ATP hydrolysis. The stoichiometry required for this function, the effect of PA700 on the lag phase of aggregation, and the temporal specificity of PA700 in this process all indicate that PA700 interacts with a subpopulation of non-native conformations that is either particularly aggregation-prone or nucleates misassociation reactions. The inhibition of off-pathway self-association reactions is also reflected in the ability of PA700 to promote refolding of citrate synthase. These results provide evidence that, in addition to binding polyubiquitin chains, PA700 contains a site(s) that recognizes and interacts with misfolded or partially denatured polypeptides. This feature supplies an additional level of substrate specificity to the 26 S proteasome and a means by which substrates are maintained in a soluble state until refolding or degradation is complete.
Strickland
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E
Program in Molecular Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75235, USA.
Hakala
K
K
Thomas
P J
PJ
DeMartino
G N
GN
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002950:Citrate (si)-Synthase / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000737:chemistry / Q000201:enzymology; D007328:Insulin / Q000378:metabolism; D008954:Models, Biological; D010447:Peptide Hydrolases / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000494:pharmacology*; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
D007328:Insulin; C085834:PA700 proteasome activator; D011506:Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate; D002950:Citrate (si)-Synthase; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease