88_22_16 The Ubiquitin/Protease system

Question 1

Proteolysis

Answer 1

expressed as inactive precursors and activity is compartmentalized to avoid affecting rest of cell

Question 2

Degradation v. repair

Answer 2

similar functional structure (compartmentalized)

Question 3

Ubiquitin

Answer 3

evolutionarily very conserved, heat stable, needed for life, 1000 targeting proteins

Question 4

Proteasome

Answer 4

Degrades ubiquitinated proteins, conserved, needed for life, 32 varieties

Question 5

Two types of proteases

Answer 5

specific and ATP dependent, or non-specific and ATP-independent

Question 6

Ubiquitin structure

Answer 6

compact, surface lysines, reactive, exposed carboxy terminus, stored in a chain to protect C-terminus

Question 7

Thiolester cascade proteins

Answer 7

E1- Ub activating enzyme (ATP dependent)
E2- Ub conjugating enzyme (mediates transfer from E1 to either E3 or substrate)
E3- Ub protein ligase (isopeptide bond from Ub to protein) (e.g. Parkin -> Parkinson’s Disease)

Question 8

Ub-Chain formation

Answer 8

Binds to lysine on substrate, lysines on Ub bind to other Ub C-termini until 4-50 present with a hydrophobic stripe - promotes interaction with proteasome, not always for degradation targeting

Question 9

Eukaryotic proteasome

Answer 9

20S catalytic core- 4 stacked 7-subunit rings form a barrel… 3 hydrolytic activities, narrow entry (alphas and internal betas brought together by a chaperone Ump1, immature sequences released upon ump1 activities)

19S regulatory particle - 2 at either end of 20S to make dumbbell, bind and release multiUb-proteins, contains ATP dependent unfolding enzymes, releases free Ub post-degradation

Question 10

Proteasome regulation

Answer 10

Allosteric bite and chew (steric freedom controls)
ATP hydrolysis
Chew and spew
Combinatorial diversity

Question 11

Diseases

Answer 11

early Parkinson’s from non-functional E2

HPAV (highrisk) from altered targeting of an E2 associated protein (E6AP) (E6 causes loss of growth control)

von Hippel-Lindau, mutant E3 subunit leaves lots of free Hif-1

lots of coupled pathways (inflammation, Ub)