How Do Cells "Digest" Proteins? The Mechanism and Function of the Ubiquitin-Proteasome System

1. How Does the Ubiquitin-Proteasome System Regulate Cellular Life Activities?

The ubiquitin-proteasome system (UPS) is one of the key mechanisms responsible for selective protein degradation within cells and is extensively involved in regulating critical biological processes such as cell proliferation, differentiation, apoptosis, and DNA repair. Dysfunction of this system is closely associated with the pathogenesis of various diseases, including neurodegenerative disorders (e.g., Alzheimer's disease, Huntington's disease), multiple types of cancer, cardiovascular diseases, and respiratory diseases. In neurodegenerative diseases, endogenous or exogenous factors lead to protein misfolding, which impairs the normal function of the UPS, resulting in the accumulation of abnormal proteins and neuronal death. During tumorigenesis, the UPS can enhance cell proliferation and anti-apoptotic capabilities by promoting the degradation of tumor suppressor proteins (e.g., p53) or inhibiting the clearance of oncoproteins. In recent years, targeting the UPS has become an important direction in therapeutic strategy research.

2. What Is the Mechanism of the Ubiquitin-Proteasome Pathway?

The core components of this system include ubiquitin (Ub), ubiquitin-activating enzyme (E1), ubiquitin-carrier protein (E2), ubiquitin ligase (E3), the 26S proteasome, and deubiquitinating enzymes (DUBs). Its function primarily involves two sequential processes: ubiquitination of target proteins and subsequent proteasomal degradation.

Ubiquitin is a conserved polypeptide composed of 76 amino acids, existing in free form or covalently bound to substrates. Ubiquitination refers to the covalent attachment of the C-terminus of ubiquitin to the ε-amino group of a lysine residue or the α-amino group at the N-terminus of a target protein. This process is catalyzed by three classes of enzymes: E1, E2, and E3:

·       E1 (ubiquitin-activating enzyme): Highly conserved, activates ubiquitin in an ATP-dependent manner and forms a thioester bond with ubiquitin.

·       E2 (ubiquitin-carrier protein): Diverse in type, carries activated ubiquitin and cooperates with E3 to recognize substrates.

·       E3 (ubiquitin ligase): The most numerous and structurally diverse, directly determines substrate specificity. Major classes include HECT-domain, RING-finger, and U-box E3 ligases. E3 functions include recognizing E2 and substrates and catalyzing ubiquitin chain formation.

Ubiquitin chains linked via Lys48 target substrates for degradation by the 26S proteasome, while Lys63-linked chains are involved in non-degradative processes such as signal transduction and DNA repair. Deubiquitinating enzymes (DUBs) remove ubiquitin tags prior to degradation, enabling ubiquitin recycling.

The 26S proteasome is a multisubunit complex consisting of a 20S core particle (CP) and a 19S regulatory particle (RP). The 20S CP has a barrel-like structure composed of four stacked rings: two outer α-rings and two inner β-rings, with proteolytic activity residing in the β1, β2, and β5 subunits. The 19S RP recognizes ubiquitinated substrates, unfolds them, and translocates them into the 20S degradation chamber. This system enables highly efficient, irreversible, and selective protein degradation.

3. What Is the Therapeutic Potential of Deubiquitinating Enzyme Inhibitors?

DUB inhibitors primarily target members of the cysteine protease and metalloprotease families, such as USP7, UCHL5, USP14, and PSMD14. Small-molecule inhibitors like P5091 and P22077 inhibit USP7 and induce apoptosis in bortezomib-resistant multiple myeloma. WP-1130 and b-AP15 simultaneously inhibit multiple DUBs, demonstrating antitumor effects in models such as colon cancer and exhibiting synergistic effects with bortezomib.

4. What Are the Application Strategies for Ubiquitinating Enzyme Modulators?

Modulating the activities of E1, E2, and E3 enzymes can influence the stability and function of substrate proteins. E1 inhibitors such as PYR-41 and PYZD-4409 are still under investigation, while MLN4924, an inhibitor of NEDD8-activating enzyme, has entered clinical trials. E2 inhibitors like CC0651 allosterically inhibit CDC34 function, blocking ubiquitin transfer. Due to their substrate specificity, E3 ligases represent ideal targets. For example, MDM2-p53 interaction inhibitors such as Nutlin-3 and RITA stabilize p53 and promote tumor cell apoptosis. Related drugs have been used in the treatment of malignancies including multiple myeloma and mantle cell lymphoma.

Conclusion

The ubiquitin-proteasome system plays a central role in maintaining protein homeostasis and cellular function, and its dysregulation is closely linked to various major diseases. Currently, drug development targeting this system has achieved substantial progress in cancer therapy, including proteasome inhibitors, deubiquitinating enzyme inhibitors, and ubiquitinating enzyme modulators. Future research will continue to deepen the understanding of the regulatory mechanisms of this system and promote the development of more selective and efficient therapeutic strategies.

Product Information