Lecture 1 - Exam 4: Protein Sorting and Vesicular Trafficking Flashcards

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1
Q

The synthesis of proteins from the endoplasmic reticulum is destined for?

A

Destined for the Golgi, endosomes, lysosomes, plasma membrane, and/or secretory vessels.

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2
Q

The synthesized proteins from the ER is destined for the Golgi, endosomes, lysosomes, plasma membrane, and/or secretory vessels. What determines protein localization?

A

Signal sequence.

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3
Q

Where does the synthesis of lipids occur?

A

Smooth ER.

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4
Q

What kind of posttranslational modifications happen and where do they happen?

A

Glycosylation and attachment of lipids are modifications that occur in the ER.

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5
Q

What are the three different orientations of membrane proteins?

A

There two types that span the membrane once, but differ in whether the carboxyl (C) or amino (N) terminus is on the cytosolic side.
There is another orientation that has multiple membrane-spanning regions.

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6
Q

Describe the insertion of a membrane protein with a cleavable signal sequence.

A

Some transmembrane proteins have a normal amino terminal signa; sequence. The signal is cleaved by the signal peptidase as the polypeptide chain crosses the membrane.

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7
Q

Describe translocation of a membrane protein with a cleavable signal sequence that is inserted into the ER.

A

Translocation is inhibited by the membrane spanning domain (~25 hydrophobic a.a.’s). This opens the translocon and allows the hydrophobic transmembrane (TM) domain to move laterally into the membrane.

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8
Q

Describe the insertion of membrane proteins with internal transmembrane sequences.

A

Other proteins are inserted into the ER by internal TM sequences that are recognized by SRP, but not cleaved by the peptidase.
These internal sequences are then brought to the translocon and the ribosome pushes the rest of the a.a. chain into the lumen of the ER.

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9
Q

The orientation of the membrane proteins depends on?
What is it driven by?

A

Depends on the amino acids immediately flanking the TM domain = positively charged amino acids stay on the cytosolic side of the translocon.
This is driven by negatively charged residues near the cytosolic side of the translocon.

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10
Q

Describe the insertion of membrane proteins that span the membrane multiple times.

A

The protein is inserted into the ER by internal TM sequences that are recognized by SRP, but not cleaved by the peptidase (like membrane proteins with internal transmembrane sequences). Again, these internal sequences are brought into the translocon and the ribosome pushes the rest of the a.a. chain into the lumen. With the same protein still intact, a second transmembrane sequence is inserted into the translocon (not recognition by SRP here, only in the beginning). The ribosome pushes the rest of the a.a. chain into the CYTOSOL. A third transmembrane sequence is inserted into the translocon and the rest of the a.a. chain is pushed into the lumen.

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11
Q

Describe posttranslational insertion of a protein with a C-terminal transmembrane sequence.

A

C-terminus is not recognized by SRP because the C-terminal sequence does not exit the ribosome until translation is complete. TRC40 brings them to a different TM receptor in the ER, GET1-GET2.

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12
Q

What happens during posttranslational translocation of proteins in the ER?

A

The Hsp70 chaperones or BiPs are thought to bind the protein as it leaves the translocon and then mediates protein folding in the ER.

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13
Q

After the Hsp70 chaperones or BiPs bind the protein as it leaves the translocon, the protein is folded in the ER (mediated by the chaperone). What happens to correctly folded proteins? What about incorrectly folded proteins?

A

Correctly folded proteins can go on to the Golgi. Incorrectly folded proteins are targeted for degradation.

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14
Q

Does the ER have a reductive environment or an oxidative environment? What does this type of environment allow?

A

An oxidative environment. This allows disulfide bonds (S-S) to form via the enzyme protein disulfide isomerase (PDI).

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15
Q

Does the cytosol have a reductive environment or an oxidative environment?

A

Reductive environment.

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16
Q

What is O-glycosylation? What is N-glycosylation?

A

O-glycosylation: The attachment of a sugar to the hydroxyl of serine (Ser, S), threonine (Thr, T).
N-glycosylation: The attachment of a sugar to the amide group of asparagine (Asn, N).

17
Q

What are the predominant sugars found in glycoproteins?

A

N-acetylgalactosamine (GalNAc) and N-acetylglucosamine (GlcNAc)

18
Q

Describe the structure of N-glycosylation (Asparagine).

A

Core group of two N-acetylglucosamines (GlcNAc) and three mannoses (complex sugars).

19
Q

Describe the structure of O-glycosylation (Serine, Threonine).

A

Core group of one N-acetylgalactosamine (GalNAc).

20
Q

Remember only specific Asn (N) residues are glycosylated. What are those residues?

A

Asn – X – Ser
or
Asn – X – Thr

X is any amino acid except proline.

21
Q

Addition of GPI anchors:
What are glycosylphosphatidylinositol (GPI) anchors?

A

GPI anchors are assembled in the ER. They contain two fatty acid chains linked to an inositol head group and an oligosaccharide portion consisting of mannose and glucosamine residues.

22
Q

Addition of GPI anchors:
GPI anchors have an oligosaccharide portion consisting of mannose and glucosamine residues. What are these added to?

A

These are added to polypeptides anchored in the membrane by a C-terminal membrane-spanning region.**

23
Q

Addition of GPI anchors:
GPI anchors have an oligosaccharide portion consisting of mannose and glucosamine residues that are added to polypeptides anchored in the membrane by a C-terminal membrane-spanning region. What happens after that?

A

The membrane-spanning region is cleaved and the new carboxy terminus is joined to the NH2 group of ethanolamine, leaving the protein attached to the membrane by the GPI anchor.

24
Q

Describe the Quality control in the ER (Calnexin-Calreticulin pathway).

A
  1. Glycoprotein exits the translocon, two glucose residues are removed.
  2. Calnexin (or calreticulin*) recognizes the glycoprotein and remove another glucose.
  3. Protein folding sensor. It will monitor hydrophobic regions.
    -If there are no problems:
    Protein will pass the quality control and will exit the ER.
    -If there are some problems:
    The folding sensor will add back a glucose residue, allowing the protein to bind back to Calnexin to attempt correcting the folding.
    -Several issues and the protein stayed too long:
    It will be recognized by EDEM1, which removes mannose residues. This prevents the protein to bind calnexin and target protein to a pathway of degradation (proteasome).
25
Q

If an excess of unfolded proteins accumulates what happens?

A

A signaling pathway called the unfolded protein response (UPR) is activated. ER-associated degradation (ERAD).

26
Q

If there’s an excess of unfolded proteins accumulates, the unfolded protein response is activated. What does this lead to?

A

It leads to expansion of the ER and production of more chaperone proteins.

27
Q

The Unfolded Protein Response:
If protein folding can’t be adjusted to a normal level, what happens?

A

The cell undergoes programmed cell death (apoptosis).

28
Q

What are the 3 receptors that the UPR activates in the ER?

A

IRE1, ATF6, and PERK

29
Q

What does the activation of IRE1, ATF6, and PERK cause?

A

Activation of these receptors causes transcription of genes encoding chaperones lipid synthesis enzymes, and ERAD proteins.

30
Q

Draw the UPR receptor actions

A

Draw them so you can remember! Slide 58

31
Q

With the PERK receptor, what happens to eIF2?

A

eIF2 is phosphorylated by PERK and therefore inhibits eIF2. When active, eIF2 brings the tRNA to the ribosome. So, that will not happen. Translation is then “dampened down” and trying to prevent too many folded proteins.