Lecture 14: Protein Degradation and Autophagy

Question 1

Why degrade proteins?

Answer 1

Incomplete or mis-sense proteins 
Cellular errors including incorporation of inaccurate amino acids or analogues 
Proteins with disruptive mutations
Products of premature termination
Products of proteolytic cleavage

Post-synthetic damage (up to 30% of newly made protein)
Mis-folded proteins
Protein ageing e.g. Oxygen free radical damage
Denaturation e.g. by heat shock

Unwanted proteins
Inactive or ‘used’ proteins
Free subunits of multimeric complexes
Other proteins made in excess

Any protein in the wrong place at the wrong time

Question 2

What are the intracellular protein degradation routes?

Answer 2

Two sites for protein degradation:

A cytosolic structure called the proteasome
OR
a membrane bound organelle called the lysosome
Membrane-bound proteins and lumenal proteins that enter via endocytosis go to lysosome

Question 3

Describe the role of ubiquitin in protein degradation

Answer 3

Ubiquitin targets proteins for degradation at the proteasome and lysosome
Essentially there are three pathways to protein degradation: Proteosome, endocytic delivery to the lysosome; autophagic delivery to the lysosome. Ubiquitin can be involved in all cases
Ubiquitin can be added to soluble proteins, ubiquitination sends it to proteasome
Receptor can be taken into endosome and ubiquitinated
Damaged mitochondria can be ubiquitinated and taken into autophagosome - process of autophagy and delivery to lysosome

Question 4

What is ubiquitin?

Answer 4

76 amino-acid polypeptide that labels proteins
Added to lysine side chains of proteins
Ubiquitination is the addition of ubiquitin to proteins

Question 5

Describe how ubiquitination takes place

Answer 5

Ubiquitin Conjugating Enzymes Add Ubiquitin to Proteins
Ubiquitin modification is an ATP-dependent process carried out by three classes of enzymes. A “ubiquitin activating enzyme” (E1), a “ubiquitin conjugating enzyme” (E2), followed by a “ubiquitin ligase” (E3)

The reactions are sequential

Question 6

Describe how ubiquitinylation is reversible

Answer 6

Ubiquitin is removed from proteins by de-ubiquitinases (DUBs) and recycled. It is generally not degraded
Dub used to break down ubiquitin chains into single ubiquitin molecule
Ubiquitin generally not degraded with the target protein- removed first, tends to be recycled

Question 7

What is autophagy?

Answer 7

Autophagy is a process of self-cannibalisation.
Cells capture their own cytoplasm and organelles and deliver them to the lysosome for nutrient release.
Selective autophagy often captures ubiquitinated cargo
Stress and starvation stimulate autophagy
Exercise produces free radicals which also cause autophagy
Generally known as starvation response - start undergoing autophagy if low in nutrients.
Double membrane forms - completely surrounds contents (organelles and cytoplasm. Deliver resulting autophagosome to lysosome and degrade the contents