II. Post-transcription | 30. Definition of proteostasis; types of intracellular protein degradation Flashcards
I. Basics
1. What is Proteostasis?
Proteostasis describes protein homeostasis and is the process that regulates proteins within the cell, in order to maintain the health of cellular proteome (entire set of proteins expressed by a genome) and the organism itself.
I. Basics
2. What is normal proteostasis due to? What happen if there is any disturbance?
- Normal proteostasis is due to the balance of many factors, such as synthesis, folding, trafficking, aggregation and normal degradation.
- If any of these are disturbed (due to mutation, aging, cell stress, injury), a variety of disorders can occur.
I. Basics
3. What is Proteostasis network?
an integrated organization of mechanisms (proteins) that control and support the genesis, folding, trafficking and turnover of proteins within and outside the cell
II. Mechanisms by which proteostasis is ensured
1. What are the 3 Mechanisms by which proteostasis is ensured?
- Regulated protein translation
- Chaperone assisted protein folding
- Protein degradation pathways.
II. Mechanisms by which proteostasis is ensured
2. What are the features of “Regulated protein translation” mechanism?
- The synthesis of a new peptide chain using the ribosome is very slow and the ribosome can even be stalled when it encounters a rare codon
- These pauses provide an opportunity for an individual protein domain to become
folded before the production of following domains
=> This facilitates the correct folding of multi-domain proteins
II. Mechanisms by which proteostasis is ensured
3A. What are the features of Chaperone assisted protein folding?
- In order to maintain proteostasis, cell uses molecular chaperons, which also aid in the assembly of proteins
- They recognize exposed segments of hydrophobic amino acids in the growing peptide chain and work to promote the proper formation of interactions that lead to the desired folded state
- Chaperonins (special class of chaperones) promote native state folding by cyclically encapsulating the peptide chain.
-> All chaperonins exhibit an open and close state, between which they can cycle. This cycling process helps to avoid undesired interactions, therefore important during folding
II. Mechanisms by which proteostasis is ensured
3B. What is the role of chaperones?
- In order to maintain proteostasis, cell uses molecular chaperons, which also aid in the assembly of proteins
- They recognize exposed segments of hydrophobic amino acids in the growing peptide chain and work to promote the proper formation of interactions that lead to the desired folded state
II. Mechanisms by which proteostasis is ensured
3C. What is the role of Chaperonins?
- Chaperonins (special class of chaperones) promote native state folding by cyclically encapsulating the peptide chain.
- All chaperonins exhibit an open and close state, between which they can cycle.
- This cycling process helps to avoid undesired interactions, therefore important during folding
II. Mechanisms by which proteostasis is ensured
4. What are the features of Protein degradation pathways?
- Protein degradation occurs in proteostasis when the cellular signals indicate the need to decrease overall cellular protein levels
- Multiple substrates are targets for proteostatic degradation: non-functional proteins, misfolded/unfolded proteins, aggregated proteins (amyloids) and proteins no longer needed to carry out cellular function
II. Mechanisms by which proteostasis is ensured
5. Why do we need to adjust each of these Mechanisms by which proteostasis is ensured?
Adjusting each of these mechanisms to the demand for proteins is essential to maintain all cellular functions relying on a correctly folded proteome
III. Proteolysis
1. What is proteolysis?
- Proteolysis is the breakdown of proteins, via hydrolytic cleavage of the peptide bond, into smaller polypeptides and amino acids.
- Proteases are the typical catalyst in these types of degradations, since protein degradations themselves are extremely slow – can also be done by other homeostatic effects, like change in temperature and pH.
III. Proteolysis
2. What are proteases?
Proteases are the typical catalyst in these types of degradations, since protein degradations themselves are extremely slow – can also be done by other homeostatic effects, like change in temperature and pH.
III. Proteolysis - Intracellular protein degradation
3. What is Intracellular protein degradation?
Intracellular degradation of proteins is the mechanism when proteins are broken down into amino acidseffects, like change in temperature and pH.
III. Proteolysis
4. By the mechanism of Intracellular degradation of proteins, what is it possible to do?
- By this mechanism it is possible to (1) remove damaged + abnormal proteins and prevent their accumulation, and (2) regulate cellular processes by removing enzymes + regulatory proteins that are no longer needed for the cell = inactivate them.
- The amino acids may then be reused for protein synthesis -> recycling
III. Proteolysis
5. What is the role of Exopeptidase?
cleaves the N- or C terminus (amino- / carboxy-peptidase)
III. Proteolysis
6. What is the role of Endopeptidase (protease)?
cleaves internal peptide bonds
III. Proteolysis
7. What are the 2 types of Intracellular protein degradation?
(1) limited proteolysis
(2) complete degradation
III. Proteolysis - limited proteolysis
8A. What is limited proteolysis?
Limited proteolysis is when the proteolysis of a polypeptide occurs after translation in protein synthesis = posttranslational modification (irreversible)
III. Proteolysis - limited proteolysis
8B. What are the 3 steps of limited proteolysis?
- Removal of N-terminal methionine
- Removal of signal sequence
- Cleavage of precursor proteins
III. Proteolysis - limited proteolysis
8C. How does Removal of N-terminal methionine occur in limited proteolysis?
- the initiating methionine may be removed during translation of the protein, so the translation never happens
- In both pro- and eukaryotes, this N-terminal may determine the half-life of the protein according to the N-end rule (= rate of protein degradation through recognition of the N-terminal residues of proteins)
III. Proteolysis - limited proteolysis
8D. How does Removal of signal sequence occur in limited proteolysis?
- N-terminal signal peptide on the protein will determine where the proteins will be active = signal will determine the final destination
- This signal peptide is removed by proteolysis after their transport through a membrane
III. Proteolysis - limited proteolysis
8E. How does Cleavage of precursor proteins occur in limited proteolysis?
- Proteins that are synthesized in the form of their precursors are cleaved to form their
final active structure - Ex: proteases are synthesized in their inactive form, so they can be safely stored in cells and are ready for release when needed. This is to ensure that they are only activated in correct location, as inappropriate activation can be very destructive for an organism
=> Proteolysis can therefore be a method of regulating biological processes by activating / inactivating proteins
III. Proteolysis - complete degradation
9. What is complete degradation?
Complete degradation is when the whole protein gets degraded at multiple sites, because it has either (1) become non-functional, (2) completed its cell cycle or (3) not needed anymore.
III. Proteolysis - complete degradation
10. What are the 2 way that complete degradation can be achieved?
- Proteolysis in lysosome
- Ubiquitin-mediated protein degradation
III. Proteolysis - complete degradation
11. What are the features of Proteolysis in lysosome?
- Lysosomes contain enzymes that are able to break down not only peptides, but other biomolecules as well. This is due to the low pH (4 - 4,5) in the interior.
- The autophagy- lysosomal pathway is a non-selective process, but it may become selective upon starvation, whereby proteins are selectively broken down.
- Lysosomes contain a large number of proteases (like cathepsins, which are hydrolytic enzymes)
III. Proteolysis - complete degradation
12. What are the features of Ubiquitin-mediated protein degradation?
- Essential biological mechanism in all living cells is the ubiquitin-proteasome pathway (UPP). Responsible for recognition, tagging and transport for degradation.
- Only proteins that are defective or need to be removed should be degraded.
III. Proteolysis - Ubiquitin-mediated protein degradation
13. What are the features of ubiquitin (Ub)?
The proteins that are defective or need to be removed should be degraded.
=> These proteins will therefore be marked with a 76 amino acid long polypeptide chain called ubiquitin (Ub).
=> This ‘’polyubiquitin tail’’ marks the protein for destruction in the proteasome.
III. Proteolysis - Ubiquitin-mediated protein degradation
14. How do Ub molecules work?
Ub molecules accumulate on the lysine residues of the target protein. Ub-tails are attached by non-α-peptide (isopeptide) bonds between carboxyl terminal of Ub and the amino groups of lysine in the target protein
- NOTE: Ubiquitin marks cytosolic proteins for degradation in proteasomes
III. Proteolysis - Ubiquitin-mediated protein degradation
15. Which enzymes are required in ubiquitin cascade?
Involves 3 enzymes:
- E1: ubiquitin activating enzyme
- E2: ubiquitin-conjugating enzyme
- E3: ubiquitin ligase
III. Proteolysis - Ubiquitin-mediated protein degradation
15. What are the 3 steps of ubiquitin cascade?
1) Activation
2) Conjugation
3) Ligation
III. Proteolysis - Ubiquitin-mediated protein degradation
15A. In ubiquitin cascade, the 1st step is ACTIVATION
=> Describe this step
- Activation of E1 by the addition of an ubiquitin molecule
- The C-terminus of ubiquitin is linked via a high-energy thioester bond to a cysteine side chain on the E1 protein.
=> This reaction requires ATP, and proceeds via a covalent AMP-ubiquitin intermediate
III. Proteolysis - Ubiquitin-mediated protein degradation
15B. In ubiquitin cascade, the 2nd step is Conjugation
=> Describe this step
- E2 catalyzes the transfer of the ubiquitin from E1 to the active site of E2 via thio-esterification reaction
- In order to perform this reaction, E2 binds to both E1 and activated ubiquitin
III. Proteolysis - Ubiquitin-mediated protein degradation
15C. In ubiquitin cascade, the 3rd or final step is Ligation
=> Describe this step
- Catalyzes the final step for the ubiquitin cascade
- Create an iso-peptide bond between the carboxyl group of the C-terminal glycine on ubiquitin and the amino group of side chain of the lysine of the target protein
- Iso-peptide bond = links a side chain amino group, rather than the α-amino group to a carboxyl group
=> When the protein is attached to an ubiquitin tag, it is targeted to the proteasome. The ubiquitin is then released and reused, while the targeted protein is degraded
