Proteins III Flashcards
Where does degradation of dietary protein occur (inside/outside cell)? Is it specific or non-specific?
Extracellular environment, non-specific
Why is proteolysis important?
- Breakdown digested protein
- Release free amino acids needed for protein synthesis
- Activation of pro-enzymes
- Degradation of regulatory proteins
- Degrade damaged/malfunctioning proteins
- Degrade foreign proteins
- Degrade mislocalized proteins
- Degrade protein made in excess
What is a chaperone? Do they require energy? Do they always work?
Ex: Hsp70 and Hsp40
Small proteins that bind to a protein while it is being translated and help it to fold correctly. Utilize ATP. It is possible for the protein to still be misfolded after chaperones have acted on it. If so, the protein could engage with a chaperonine.
What is a chaperonine?
Do they require energy?
Do they always work?
Are these specific or non-specific folding mechanims?
Ex: Hsp60
Barrel like protein that acts on misfolded proteins only.
Requires ATP to fold protein correctly.
The protein can still be misfolded after the chaperonine has acted upon it. If this were the case, then the protein would be targeted for degraation.
Non-specific.
Ubiquitin
- What is it’s job?
- Describe the amino acids that are relevant to it’s structure.
- Ubiquitin’s job is to find the misfolded substrate and attach to it and then recruit more ubiquitins to attach
- Seven lysine residues on surface and highly reactive glycine at carboxy terminus that is accessible for reaction. Reactive glycine binds to specific lysine on target protein.
Describe the expression of ubiquitin.
It is always expressed as a fusion protein i.e. multiple Ub monomers are translated into a single protein, which is then cleaved to release the invididual monomers that ubiquitinylate proteins.
There are 3 enzymes involved in ubiquitinylation. What are they (general terms)?
E1: Activates Ub
E2: Conjugates Ub to protein to either E3 or the substrate
E3: Attaches Ub to substrate
What kind of bond is made between Ub and it’s target proteins?
Isopeptide, very stable
Multiple Ubs attached to one protein is called poly-Ub.
- What is the effect of Poly-Ub?
- How is this accomplished?
- Each Ub added to a Ub chain amplifies the signal to the cell that the protein needs to be degraded
- Accomplishes this b/c Ubs have regions of hydrophobic patches, when poly-Ub, these hydrophobic areas line up to form a hydrophobic stripe, which is the primary determinant for promoting interaction with proteasome
Is ubiquitinylation always a signal for degradation?
No - it depends on the way that the bonds between Ub and the protein and other Ubs in Poly-Ub are made. Some Ub sites signal degradation, others do not.
- Mono-Ub?
- Multi-Ub?
- Poly - Ub?
Explain how Ubiquitin is able to recognize so many different sites on different proteins?
Ubiquitin displays combinatorial diversity at the level of the enzymes that control its function. There are different kinds of E2 and E3 enzymes. Additionally, E3 is a multi-subunit enzyme. As such, the subunits can change and that will change the signaling pathways that it is involved in. Thus, different combinations of E2 and E3 result in the ability to degrade different substances
The proteasome can be broken down into three subunits: 19S –> 20S –> 19S. What is the function of the 19S and 20S subunits?
20S - houses hydrolytic residues
19S - Bind to Ub via recognition of hydrophobic stripe, contains unfolding enzymes that begin to denature protein, funnels protein into 20S subunit
Which steps of protein degradation require energy?
Which steps do not?
How does the cell regulate the number of proteasomes in it?
TORC 1 = master growth controller, adjusts proteasome levels in response to cellular need, inhibition = increased amount of proteasome
