Lecture 14 - Protein Degredation and Autophagy Flashcards
Three instances when proteins need to be degraded?
- Incomplete/mis-sense.
- Damaged.
- Unwanted.
What does mis-sense mean?
Proteins not required.
When do incomplete/missense proteins occur?
- Cellular errors, e.g. incorrect AA sequence.
- Proteins with disulphide mutations.
- Products of premature termination.
Why do damaged proteins occur?
- Mis-folded.
- Protein ageing.
- Denaturation.
Why can some proteins be unwanted?
- Inactive/’used’
- Excess of a specific subunit.
- Made in excess.
There are two degradation routes. Name them.
Proteosome and lysosome.
Difference between a proteosome and a lysosome?
Proteosome = cytosolic structure, soluble proteins degraded here. Lysosome = membrane-bound organelle, membrane proteins (e.g. those that enter by endocytosis), autophagy involved.
How are proteins targeted to the lysosome and proteosome for degradation?
Ubiquitin.
Structure of ubiquitin?
76 AA polypeptide, mainly added to lysine side chains.
Ubiquitin is added to proteins via 3 Ub enzymes. Name them.
E1, E2 and E3.
Function of the 3 Ub enzymes?
E1 = activates Ub. ATP-dependent. Ub now bound to E1. E2 = receives Ub from E1 and either uses E3 Ub ligase to transfer the Ub directly to the substrate, or uses an E3 HECT domain.
E3 either has a ring domain or a HECT domain.
Difference between a ring domain and a HECT domain?
Ring domain = ligase; Ub goes directly from E2 –> substrate.
HECT domain = goes from E2, to E3, to the substrate.
Two types of ubiquitination?
Mono- or poly-
Ub molecules attach to lysine resides via?
Isopeptide linkage.
Explain the different types of polyubiquitination.
Can either have a chain structure or can be branched. Branched structure forms because Ub molecules have seven lysine residues.
Is ubiquitination reversible? Explain your answer.
YES. De-ubiquitinases (DUBs) can remove Ubs from proteins and recycle them.
How are different Ub proteins targeted to different regions of degradation?
Different Ub conformations = different molecular tags.
Proteosome uses two types of receptor for protein degradation. Name them.
Intrinsic receptor = possesses Ub binding domains to bind ubiquitinated cargo. Bound to the proteosome.
Shuttling receptor = Ub binding domain AND a proteosome protein binding domain.
The lysosome can perform autophagy and endocytosis which both use a different type of receptor. Name them.
Autophagy - receptor with a Ub binding domain and a domain that targets it to the autophagosome membrane.
Endocytosis - two domains, one targets the membrane and the other binds Ub.
Structure of the proteosome?
Central cylinder and cap proteins.
Central cylinder contains the proteases. Ub protein is targeted to the cap proteins where ATP is used to unfold the protein. Protein then spread through the proteosome where it is divided into its AA’s.
Structure of cap proteins?
Hexameric AAA-ATPase structure.
An example of proteosome-mediated degradation: the tumour suppressor protein p53.
- p53 is ubiquitinated.
- p53 causes the expression of Mdm2, an E3-Ub ligase.
- p53 is polyubiquitinated and targeted to the proteosome.
- Mdm2 levels decrease. Autoregulatory.
- MdmX inhibits transcription of p53. Mdm2 regulatesthe levels of Mdmx via ubiquitination.
When receptors enter an early endosome for degradation in a lysosome, they can techically still signal. ALL PARTS OF THE RECEPTOR MUST BE INTERNALISED. How does this occur?
Invaginations of the endosomal membrane to form a multivescular body (MVB).
Which family of proteins are responsible for the formation of MVBs?
ESCRT proteins.
ESCRT = Endosomal Sorting Complexes Required for Transport.
In simple terms, what is autophagy?
Self eating. Cells completely surround an organelle/part of their cytoplasm using a membrane and deliver
How is autophagy stimulated?
Stress, starvation and exercise.
Types of autophagy?
- Macroautophagy
- Mitophagy - degredation of mitochondria.
- Pexophagy - perioxisomes.
Autophagy regulates energy homeostasis. Explain this.
Since autophagy is a starvation response, material is degraded to its building blocks to provide energy.
What happens if autophagy is impaired?
- Lack of nutrients.
- Damaged mitochondria cannot be removed - ROS accumulation.
- Bacterial infection cannot be cleared = chronic infection.
