Proteasome channel opening as a rate-limiting step in the ubiquitin-proteasome system

The 26S proteasome eliminates multiubiquitinated proteins in cytosol and nucleus, and from the secretory pathway by a mechanism known as ER-associated degradation (ERAD). Access to the proteasomal 20S catalytic core particle is hindered by conserved N-terminal tails of α-subunits that form a gated pore into the central channel. Hence, the isolated 20S core particle possesses slower peptide hydrolysis rates and cannot degrade multiubiquitinated proteins. Purified catalytic particles from an α3α7ΔN open channel double mutant, in which the N-terminal tails of α-subunits from opposite sites of the α ring are deleted, showed significantly enhanced peptidase activity and proteolytic properties. Here we show that also in vivo the access of substrates to the proteasomal catalytic chamber partially limits the overall rate of protein elimination. This regulation applies to unstable cytosolic proteins of the N-end rule and ubiquitin fusion degradation (UFTD) pathways, as well as to ERAD substrates that must dislocate from the ER back to the cytosol in order to become ubiquitinated and degraded by the proteasome. Hence, even for a complicated multistep process such as ERAD, traffic through the proteasome itself is partially rate limiting for the entire proteolytic process. However, proteasome gating can be added to a growing list of phenomena that distinguish membrane ERAD substrates from lumenal ones because while gating hinders access of lumenal substrates, it is less effective in controlling the entry of membrane substrates. The open channel mutant is a new class of proteasome mutant, which is unrelated to the catalytic protease active sites or to the "classical" regulatory particle mutants. Its improved performance at high temperatures is in stark contrast to the behavior of the "classical" mutants, suggesting that the α3α7ΔN mutant adapts better to mild stress conditions.