Quality Control of Proteins Flashcards
Understand protein folding
Protein folding is the process by which a linear chain of amino acids (polypeptide) acquires its functional three-dimensional structure
Must fold correctly to perform their biological roles
The folding process is typically spontaneous, driven by the physical and chemical properties of the amino acids and their interactions with each other and the surrounding environment
Describe chaperones
Bind to newly synthesised polypeptides preventing premature folding
Prevent aggregation - prevent hydrophobic regions from clumping together by binding to them, keeping the protein in a foldable state
Refolding of mis-folded proteins
Stimulate protein degradation
Describe the importance of protein folding:
Functionality - protein’s shape determines its function
Specificity - Correct folding ensures that proteins interact specifically with their target molecules
Stability - Properly folded proteins are more stable and less likely to degrade
Describe HSP70 proteins:
Heat shock protein
ATP bound HSP70 binds to hydrophobic regions of the unfolded protein
ATP is hydrolysed to ADP+Pi Causes conformational change and tighter HSP70 binding
HSP40 disassociates and another ATP binds to remove HSP70 from the protein
Protect cells from heat induced stress preventing protein misfolding
Describe the unfolded protein response :
Signaling system that detects misfolded proteins within the endoplasmic reticulum and coordinates a cellular response that aims to restore protein homeostasis
UPR is initiated by ER stress
The UPR in humans is known to be composed of 3 distinct arms (ATF6, PERK and IRE1) which promote:
Short term – reduction of protein production / misfolding
Long term – increased ER capacity
If the UPR remains active for a prolonged period the cell undergoes apoptosis to prevent the retention of damaged/rouge cells
Describe the IRE1 pathway
Inositol-Requiring Enzyme 1
A protein kinase and UPR sensor that monitors the status of unfolded protein inside the ER lumen.
Also acts as an endonuclease that cleaves RNAs in the middle
Describe the PERK pathway:
Protein kinase-like Endoplasmic Reticulum Kinase
Kinase which phosphorylates translation initiating factors (eIF2α) to prevent translation.
Leads to general shutdown of mRNA translation machinery but selectively activates translation of proteins that contribute to stress response especially ER chaperones.
Describe the ATF6 pathway:
Activating Transcription Factor 6
Transcription factor kept anchored to the ER membrane. Misfolded proteins activate proteases which cleave ATF6 from the ER and it enters nucleus to induce proteins that contribute to stress response, including ER chaperones
Describe HSP60 proteins:
HSP60 proteins exist as a ‘large barrel complex’, mainly reside in Mitochondria
Act after protein has been fully synthesised
Hydrophobic regions of misfolded proteins are captured by rim of the HSP60 barrel
ATP and ‘protein cap’ are added and protein now in confined ‘isolation chamber’ allowed to refold
After a defined time (15 secs) ATP hydrolysis occurs and complex is weakened
More ATP binds to eject protein folded or not, cycle repeated
Known as chaperonins
HSP60 helps refold damaged proteins to prevent their aggregation during heat induced stress
What is the ubiquitin system ?
The ubiquitin system is a cellular process that tags proteins for degradation using a small protein called ubiquitin
Tagging occurs through a series of enzymatic steps involving three main enzyme - E1, E2, E3
Once a protein is marked with ubiquitin, it is recognized by the proteasome, a large protein complex that breaks down the tagged protein into smaller peptides
Describe -
E1
E2
E3
E1 - Ubiquitin activating enzyme
Hydrolysis of ATP
Carboxyl end of ubiquitin connected via thioster bond
E2 - Ubiquitin conjugating enzyme
Activated Ubiquitin transferred to cysteine residue
E3 - Ubiquitin protein ligation
Aids in transfer of the Ubiquitin onto target protein
Describe the process of ubiquitination:
Ubiquitin is added to target protein lysine residues via an enzymatic process called ubiquitination
Ubiquitination results in isopeptide bonds between the c-terminal glycine of ubiquitin and the lysine of the target protein
This process controls levels of short-lived proteins and unfolded proteins
What are the different types of ubiquitination ?
Monoubiquitination: addition 1 ubiquitin to a single lysine residue
Multi-ubiquitination: addition of multiple ubiquitin molecules to multiple different lysine residues
Poly-ubiquitination: addition of a ubiquitin chain (4+) to a single lysine residue
What happens when Chaperone / HSP system
fails or is overwhelmed?
Activation of the Unfolded Protein Response (UPR)
Activation of the proteolytic system
Describe the structure of proteasome
Proteasesome is a multi-protein complex responsible for degrading damaged or unneeded proteins into smaller peptides.
* The 26S proteasome is the most common form, composed of two main structures: the 20S core particle (CP) and one or two 19S regulatory particles (RP).
* The 20S core particle is a barrel-shaped, cylindrical complex with 28 subunits arranged in four stacked heptameric rings.
* The 19S regulatory particle is located at one or both ends of the 20S core particle and consists of two subcomplexes: the Base Subcomplex and the Lid Subcomplex.
