Regulated Protein Destruction

Question 1

zymogen

Answer 1

inactive enzyme

Question 2

ubiquitin

Answer 2

all euks + mark for destruction

Question 3

why proteolysis

Answer 3

mislocalized, stoic excess, degrade reg/ damage, activate proteins, etc

Question 4

proteolysis enzyme traits

Answer 4

1. expressed as inactive precursor

2. Compartmentalized

Question 5

regulations of proteolysis

Answer 5

proenzymes, compartmentalization, pH, substrate induced

Question 6

Proenzyme

Answer 6

pro attached to enzyme that inactivates-remove pro to activate

Question 7

where do substrates come from

Answer 7

post translational protein damage, coo translation damge (ribosome makes junk), regulatory proteins (undamaged but unncessary)

Question 8

Ub/Proteasome degradation +regulatory pathwayh

Answer 8

used to regualate important reg pathway proteins

Question 9

ub/proteasome pathway

Answer 9

no mutations available (death if there is mut), multiple genes for ubi but only 1 type of ubi protein, 1000 proteins that target this pathway (may have mutations)

Question 10

Two types of proteases

Answer 10

Specific (ATP DEPENDENT) and nonspecific (no ATP needed-exothermic)-attack any conspicuous protein (not normally seen)

Question 11

Ub structure

Answer 11

Reactive carboxy terminus, 7 lysines on surface, several hydrophobic patches on surface, fist with thumb extended

Question 12

Ub expression

Answer 12

Fusion protein, C terminuses blocked, ub Carboxyl terminal hydrase-cut blockages (blockages are how ub are attached together)

Question 13

ub Carboxyl terminal hydrase

Answer 13

cut blockages between newly synthesized ub

Question 14

3 enzymes for ub to substrate

Answer 14

Ub activating enzyme, ub conjcuating enzyme, ub-protein ligase

Question 15

Thiolester cascade (probably draw this)

Answer 15

1. ub activating enzyme has cystene residue attached to sulfhydrl (SH) group
2. adenylate Ub with ATP (???? you can do this)
3. ub makes thioester bond (not very stable) with S (H leaves)
4. Ub conj enzyme then takes ub
5. then goes to ub protein ligase
6. ub binds to substrate with isopeptide bond
7. attach next ub to first using lysine residue and isopeptide bond
8. hydrophobic patches collapse=hydrophobic strip

Question 16

Isopeptdie bond

Answer 16

how ub binds to protein-very stable and reversible

Question 17

multiub for cell targeting

Answer 17

residues bind to distil end lysine-can not collapse

Question 18

Combinatorial diversity

Answer 18

1000’s of substrates need to be destroyed but only so many proteasomes-use different e2 or e3 (e1 always stays same)

Question 19

Two particles of euk proteasome

Answer 19

20S catalytic cofe and 19S regulatory core combination of 20 and 19 is spontaneous…finished product is 20 S proteasome

Question 20

20S core

Answer 20

spotaneus 7 subunit barrel, beta units (only 3 have hydrolytic activity), also alpha small hole too

Question 21

19S regulatory protein

Answer 21

can bind multi ub proteins+unfold enzymes with ATP-6 ATPases, feeds unfolded protein through small hole, ub is recycled as go in

Question 22

Regulatory methods in proteasome (4)

Answer 22

1. Allosteric-big in, holds up other big going in, degraded, other big can go in
2. unfold enzymes with ATP-6 ATPases, feeds unfolded protein through small hole
3. Recycling and peptide release-lots of stuff goes in, then degrade, then spew small parts out
4. Cominbatory (swtiching subunits to affect desired protein)

Question 23

Proteasome and neuro junctions

Answer 23

When synapse unused, proteasome degrades

Question 24

ATPuses

Answer 24

Substrate unfodling, maintain stability, start of ubiquination

Question 25

Parkinson disease caused by

Answer 25

mutation in e3 ligase

Question 26

hect 3 ligase

Answer 26

E6AP (other name) catalyzes many proteins, HPV binds to complex and makes enzyme degrade p53

Question 27

HIF-1 (e3 protein is 3 sununits)

Answer 27

required for activation genes that respond under conditions of hypoxia-change subunit and degrade different protein

Question 28

VHL syndrome

Answer 28

HIF-1 (1 subunit difference)-e3 ligase can no longer reocgnize substrate

Question 29

Stress induced NFkappa pathway (draw)

Answer 29

1. NFk precursor (p-65) processed by proteasome +meets with other protein (P-50) in cytosol-becomes a TF
2. Complex inactivated by IkappaBalpha
3. Stress-activates inactive Ikappa kinase which phosphorylate IkappaBalpha and inactivates
4. TF can go to nuceleus
5. One gene product is IkappaBalpha-negatie feedback

Question 30

3 events that need proteasome in NFkappa pathway

Answer 30

1. Maturation of p-65
2. Degradation of inhibitor
3. Activation of kinase

Question 31

Immunoproteasome

Answer 31

only 3 subunits have hydrolytic (instead of 7), creates longer peptides with specific hydrophobitiy, these fit in cleft in MHCI molecules and go to cell surface and display antigen
-essentially fragments are generated differently (cuz they fit into cleft)

Question 32

Proteins that make noncov/cov with

Answer 32

cov-substrates, E1, e2, e3, other ubiquitin, expressed as ub fusion protein
noncov-chain and proteasome, ubi collapse as stack