Proteins: ER & Golgi (Lect 8)

Question 1

SIgnal Hypothesis

Answer 1

(Step 1) Ribosomes unattached to ER starts making protein. If nascent peptide as “signal peptide”, then ribosome plus growing chain will attach to ER membrane, and growing chain will enter ER as it grows (co-translational import).

Question 2

How does Ribosome get to the ER?

Answer 2

a) Nascent protein chain (not rib) has LS
b) LS = SP = section of growing peptide often at amino end
c) SP binds to SRP
d) SPR temporarily blocks translation, ferries nascent protein to ER. Helps attach rib to ER so growing chain can enter pore (translocon) in ER.

Question 3

SRP

Answer 3

Signal Recognition Particle
(example of ribonucleoprotein particle - RNP)

Structure: proteins + RNA

Question 4

How does growing chain enter ER?

Answer 4

a) SRP binds to SRP receptor on ER (docking protein)
b) ER’s Translocon (gated pore/channel through membrane) allows growing chain to pass through membrane as chain is made. Pore is closed until ribosome with growing chain gets into position.

c) Ribosome/translocon complex formation occurs:
- SRP releases, recycles–GTP split
- rib binds to pore/translocon
- translocon opens & peptide enters (as loop)
- rib resumes translation

Question 5

What do you need to get into the Nucleus vs. ER?

Answer 5

Nucleus:

• Address: NLS
• Middle Man: Transporter proteins (importin)
• Surface receptor protein: Nuclear Pore Complex

ER:

• Address: SP
• Middle man: SRP
• Surface receptor protein: Docking Protein (SRP receptor)

Question 6

What happens inside the ER?

Answer 6

1. Chaperone proteins assist in protein folding. (HSP 70: binds to hydrophobic regions, Heat Shock Protein)
2. S-S Bonds are made (proteins in cyto do not have S-S bonds, but do contain cystines and have free SH groups.)
3. Start of N-glycosylation

Question 7

N-glycosylation

Answer 7

oligosaccharide s added to the N of the amide of asparagine side chains (starts in ER and ends in Golgi)

Question 8

O-glycosylation

Answer 8

oligo is added to O in hydroxyl side chains of ser or thr. Occurs in the Golgi.

Question 9

What happens to proteins in ER that do not fold properly?

Answer 9

a) transport to cytosol
b) ubiquitin added to side chains of lysine (multiple molecules mark for proteasome)
c) proteasome degrades Uq-tagged proteins to fragments at expense of ATP. (Uq gets recycled. Short peptides get put on cell surface for immune system recognition)

Question 10

Compare and contrast proteasomes and lysosomes

Answer 10

Lysosomes:

• generally degrade material from OUTSIDE cell
• single molecule of Uq
• cytoplasmic proteins and organelles can be degraded in lysosomes under certain conditions.

Question 11

What are the 2 sides of the Golgi stack called?

Answer 11

Cis/forming face (closest to ER and nucleus)

Trans/maturing face (away from nucleus)

Question 12

What are the 3 basic parts of a Golgi stack?

Answer 12

• Cis Golgi Network (may include fusing vesicles)
• medial cisterane
• Trans-Golgi network (may include budding vesicles)

Question 13

Path Overview of proteins in EMS

Answer 13

Cyto to ER membrane to ER lumen/membrane to vesicle budding off of ER to cis Golgi, meidal Golgi, trans-Golgi to destination

Question 14

What reactions take place in the Golgi?

Answer 14

• Finish N-glycosylation: oligo that was added to glycoproteins in ER is modified. (attached to N of amide side chains of asparagines
• Do O glycosylation: sugars added to “O” of the hydroxyl of the side chain of ser and thr
• Assemble sugars of proteoglycans (linear chains of repeating sequences = GAGs)
• Concentrate, sort proteins in TGN (different areas of Golgi have receptors that trap proteins going to different destinations.

Question 15

Do you expect acid phosphatase to reach lysosomes if the is an M6P-adding mutation?

Answer 15

No: acid phosphatase reaches lysosomes by a different mechanism than most hydrolyses. Does not use M6P receptors.

We know this bc ppl with I cell disease have AP in right place.