Disassembly of the self-assembled, double-ring structure of proteasome α7 homo-tetradecamer by α6

The 20S core particle of the eukaryotic proteasome is composed of two alpha- and two beta-rings, each of which is a hetero-heptamer composed of seven homologous but distinct subunits. Although formation of the eukaryotic proteasome is a highly ordered process assisted by assembly chaperones, alpha 7, an alpha-ring component, has the unique property of self-assembling into a homo-tetradecamer. We used biophysical methods to characterize the oligomeric states of this proteasome subunit and its interaction with alpha 6, which makes direct contacts with alpha 7 in the proteasome alpha-ring. We determined a crystal structure of the alpha 7 tetradecamer, which has a double-ring structure. Sedimentation velocity analytical ultracentrifugation and mass spectrometric analysis under non-denaturing conditions revealed that alpha 7 exclusively exists as homo-tetradecamer in solution and that its double-ring structure is disassembled upon the addition of alpha 6, resulting in a 1:7 hetero-octameric alpha 6-alpha 7 complex. Our findings suggest that proteasome formation involves the disassembly of non-native oligomers, which are assembly intermediates.