Ubiquitin mediated-protein degradation is one of the most important pathway in intracellular protein degradation. The discovery of this system, that been called Ubiquitin Proteasome System (UPS), was awarded a Nobel prize in 2004. It is because of their importance in orchestrating many protein degradation. Thus, will control the protein level that regulate many diverse action.
Protein undergoes degradation to maintain its dynamic level following the synthesis. So that, the protein turnover (degradation and resynthesis) must be tightly controlled to achieve the dynamic balance. Intracellular protein degradation (proteolysis) should be maintained spesifically and selectively because each Intracellular protein has different half life that vary from minutes to days. Some of them needs special signaling pathway.
There are three type of protein degradation in mammals (summarized in the table below)
type | Explanation |
Digestion of dietary protein in the gut and absorption of amino acids | This is the process of ingested protein degradation (external protein). After degraded mechanically, the proteins will be degraded enzymatically into their absorbable form (amino acids). Some protease enzymes have pivotal role in this protein degradation, such as: endoprotease that cleave protein backbone (trypsin, chymotrypsin), exopeptidase that remove residues from N-terminus to C-terminus (carboxypeptidases), and peptidase that split oligonucleotide to smaller absorbable nucleotide or amino acids. These enzymatic mechanisms occured in the different part of gut. |
Degradation of extracellular protein | It is one of degradation system for body’s own protein. It is mediated by lysosomal degradation system. This system degraded all extracellular protein and 20% of intracellular proten. Some extracellular proteins that be degraded by this pathway are: LDL, peptide hormone and cell surface membrane protein. |
Degradation of intracellular protein | Degradation of intracellular protein mainly mediated by Ubiquitin Proteasome System (UPS). Almost all intracellular protein (80%) normally degraded by this pathway, especially intracellular proteins which have short half life. |
In this lecture we will focus on intracellular degradation system.
Lysosomal degradation system
Lysosome is one of organelles within cell which is produced by golgi apparatus. Lysosome contains enzymes: carbohydrases, lipases, nucleases, and proteases. These enzymes are very important in the degradation of waste materials, cellular debris, excess or worn-out organelles and invaded microorganisms. Lysosome can digest many particles that enter cell. Some solid particles can be digested after endocytosis, for example membrane bound protein. Pinocytosis will mediate digestion of some non-specific extracellular protein. After pinocytosis, the extracellular protein will be covered by a mantle from cell membrane and then digested by lysosomal enzymes. Lysosome can digest proteins within cell by autophagy mechanism, for example digestion of warn-out organelles.
Ubiiquitin Proteasome System
Ubiquitin Proteasome System (UPS) has important role in many cell process by degrading specific intracellular protein that takes part in specific process, such as cell growth, cell differentiation, cell cycle, signal transduction etc. UPS also has function in degrading misfolded protein, mislocated protein or mutant protein. Because of its importance in many intracellular proteins, any malfunction of UPS may leads to pathological condition. For example, malfunction of UPS in degradation of tau protein that leads to Alzheimer’s disease.
Mechanism of UPS can be divided into two major process: ubiquitination and proteolysis.
1.
Ubiquitination
Ubiquitination is a process of ubiquitin conjugation to mark the spesific protein that want to be degraded. Ubiquitination process can be summarized by the figure below:
By this figure we can see that inspite of Ubiquitin (Ub), UPS also needs three important protein: E1, E2, and E3. E1 is an ubiquitin activating enzyme that will activate Ubiquitin in its C-terminus. The activated ubiquitin then will be transferred to E2 (Ubiquitin conjugating enzymes). Finally ubiquitin will be attached to target protein by E3 (ubiquitin-protein ligase). It needs polyubiquitin chain (at least 4 ubiquitin) to be transferred to target protein before these protein becomes degradable. Mammalian has so many E3 types, more than 1000, that gives them capacity to degrade many intracellular proteins. Big number of E3 types makes them very interesting to be explored.
E3 has three mechanisms to transfer ubiqitin toward target proteins. There are constitutive recognition, modification and association with an ancillary protein. Constitutie recognition allows the substrate to be recognized without any previous alteration and are degraded at the same rate of different condition. In other case, target protein (substrate) should be modificated before attach E3, for example: phosphorylation of IkBα. In the third mechanism, targeted protein should attach ancillary protein before being transferred to E3. For example, the association of p53 and HPV oncoprotein E6.
2. Proteolysis
Protein degradation in UPS needs 26S proteasome (deubiquitinating enzymes). Next Ubiquitin as a tag is going to be recycled for reubiquitination. Proteins are tightly controlled in this degradation system. They should have spesific recognition structure. Short-lived proteins have Arg, Lys, Phe, Leu, or Trp at their N-terminus. They will be degraded within 3 minitues in vivo. In the other hand, proteins that have Cys, Ala, Ser, Thr, Gly, Val, or Met is present at their N-terminus will be degraded within 30 hours. It leads them become long-live proteins. Degradation of labelled protein occured in 26 S proteasome.
26 S proteasome is assembled from core unit (20S) and regulatory unit (19S). Core unit is composed by two identical alfa ring and two outer Beta ring. Core unit function is as a catalytic center that degrade protein into small peptide followed by release of reusable ubiquitin via deubiquitination enzyme. The 19S regulatory unit is also an important part. It has several function in binding to polyubiquitinated protein, unfolding the peptide chain, opening the access port in the outer ring, and transferring the unfolded protein into catalytic center.
Beside their capacity to degrade many intracellular proteins, proteasomes also contains deubiqutination (DUB) enzymes which are key regulators for ubiquitin-mediated pathways. DUB has diverse action, such as:
- Recycling the ubiquitin.
This function is facilitated by cysteine proteses that catalyze polyubiquitine cleavage into reusable ubiquitin monomers.
- Regulation of protein degradation.
It consists of editing and reversal of ubiquitin chains.
- Ubiquitin biosynthesis.
DUB will remove ubiquitin with additional C terminal residue to expose C terminal glysine that has active site.
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