*4.2. Ubiquitin Proteasome System*

Misfolded or destabilized proteins undergo intracellular proteolysis through two main pathways: the UPS and the autophagy-lysosome pathway. More than 80% of native and misfolded intracellular proteins undergo UPS-mediated degradation [28]. The UPS functions in both the nucleus and the cytosol to recycle and degrade soluble proteins. On the other hand, autophagy functions only in the cytoplasm and generally eliminates large, insoluble aggregates and degenerated organelles that escaped from the UPS [68]. A detailed explanation of the autophagy-lysosome pathway has been provided elsewhere [78]. In this review, we focus mainly on UPS-mediated protein degradation and its association with chaperones during the regulation of PAH and FAH proteins.

Targeting a substrate protein for UPS degradation requires the covalent attachment of Ub molecule/s, which is called ubiquitination. Ubiquitination is achieved when three enzyme families work consecutively: ubiquitin-activating enzyme (E1) activates ubiquitin by ATP hydrolysis and forms a thioester link between its own cysteine residue and the C-terminal carboxyl group of Ub. Second, ubiquitin-conjugating enzyme (E2) receives Ub from E1 by a trans-thiolation reaction in which Ub binds to the cysteine residue of E2. Third, the ubiquitin-protein ligase (E3) and E2 together position the target protein substrate and attach the ubiquitin moiety to the ε-amino group in a lysine residue on the target protein. The ubiquitin-conjugated proteins are then recognized and degraded by the 26S proteasome unless the ubiquitin chains are removed by a crucial set of enzymes called deubiquitinating enzymes (DUBs) [79,80] (Figure 1).

The role of the UPS has mainly been characterized in neurodegenerative diseases [81]. In gain-of-function disorders such as Alzheimer's, Parkinson's, Huntington's, and Creutzfeldt-Jakob diseases, the misfolded proteins accumulate to form aggregates and fail to undergo proteasomal degradation because of dysfunction in the UPS. In Alzheimer's disease, for example, the amyloid precursor protein is cleaved into amyloid β (Aβ) peptides to form intraneuronal neurofibrillary tangles. The Aβ peptides inhibit UPS-mediated degradation by binding to the catalytic core of the 26S proteasome, thereby inhibiting its chymotrypsin-like activity [82]. Interestingly, the molecular chaperones Hsp70 and Hsp40 reduce the accumulation of Aβ aggregates [83]. Therefore, the UPS and molecular chaperones balance each other's functions.
