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Biochemistry > Week 10 - Protein Sorting and Intracellular Traffic > Flashcards

Week 10 - Protein Sorting and Intracellular Traffic Flashcards

1
Q

What is the rough endoplasmic reticulus?

A

A network of membrane-enclosed tubules and sacs (cristernae) that extends from the outer layer of the nuclear envelope throughout the cytoplasm.

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2
Q

What is the endoplasmic reticulus?

A

ER membrane is continuous with the outer membrane of the nuclear envelope. Consisting of vast networks of tubules which extend throughout the cytoplasm. Not completely discrete organelles.

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3
Q

What are the funtions of the smooth endoplasmic reticulus?

A

1 - synthesis of lipids eg. phospholipids and cholesterol which from cell membranes
2 - synthesis of steroid hormones
3 - storage and release of calcium from sarcoplasmic reticulum
4 - detoxification eg. alcohol

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4
Q

What are the functions of the rough endoplasmic reticulus?

A

The site of protein synthesis and processing for proteins secreted from the cell

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5
Q

What is the process where proteins are imported to the ER?

A
  • called co-translational translocation
  • one end of the protein is attached to the ribosome whilst the other end itserts itself into the ER - the protein will remain in its primary sequence, DOES NOT need a chaperone protein
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6
Q

What are chaperone proteins?

A

They bind and escort other proteins (preventing folding/damage)

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7
Q

How do signal Recognition Particles work?

A
  • SNP recognises the signal squence on the N-terminal of the growing polypeptide
  • it binds to the signal sequence and the ribosomes
  • SNP consists of RNA and 6 proteins
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8
Q

What has to occur for co-translational translocation?

A

1 - Be recognised by Signal Recognition Particle
2 - Be recognised by the ER and embedded within the ER membrane

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9
Q

Explain the process of Protein Synthesis

A
  • Ribosomes bound to ER
  • Co-translational translocation
  • Import signal to the ER
  • Recognised by SRP
  • SRP binds to SRP receptor (translocation protein)
  • goes to the ER lumen or ER membrane
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10
Q

What happens once the ribosome has been translated?

A

The mRNA is released back into the free ribosome pool
The mRNA might still remain bound to the ER as it translated by multiple ribosomes
Ultimately the mRNA will be degraded

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11
Q

What is Exocytosis?

A

When a protein is released from the cell and it is then fully transported to the ER lumen
- If a protein is destined to be a membrane protein then it will get embedded into the ER membrane however

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12
Q

What happens inside the ER?

A

Protein will be modified in 2 ways:
1- Fold into its 3D conformation via: ionic bonds, hydrogen bonds, van der waal attraction, disulphide bond formation
2 - Glycosylation

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13
Q

What is protein Glycosylation?

A
  • refers to the addition of a sugar to a protein
  • it is VERY specific to proteins that enter the ER
    1- Quality control
    2- Recognition
    3- Protection
  • a precursor oligosaccharide composed of 14 sugars is added to proteins via the N-terminius of an asparagine side chair
  • actually occurs during protein synthesis as proteins enter ER lumen
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14
Q

Explain Quality Control

A

1 - If a protein is correctly folded 3x glucose are cleaved from N-linked oligosaccharide
2 - if it doesnt fold correctly, glucosyl transferase enzymes add a single glucose back
3 - calnexin binds to unfolded protein to prevent aggregation
4- removal of the terminal glucose (enzyme called glucosidase) releases protein from calnexin
5 - glucosyl transferase determines if protein is corrctly folded. If not a single glucose is added back.

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15
Q

What do unfolded protein response do?

A

1- Inhibit protein synthesis
2- degrade misfolded proteins
3- increase trasncription of chaperones
- BiP
- Calreticulin
- Calnexin
If problem persists - apoptosis (programmed cell death) occurs

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16
Q

What does the ER receptor PERK do?

A
  • responsible for pausing translation
17
Q

What does the ER receptors IRE1 and ATF6 do?

A

protein degradation and activation of genes to increase protein folding capacity

18
Q

Describe the process of the Pulse-Chase Experiment?

A

1- Addition of radiolabelled amino acids to cell
2- All newly formed proteins are radiolabelled - pulse
3- The cells are then returned to non-radioactive medium - chase

19
Q

What are the 3 Intracellular pathways?

A

1- Secretory
2- Endocytic
3- Retrieval

20
Q

What are the types of vesicles used to transport proteins?

A
  • COP 2 = from ER
  • COP1 = from golgi
  • Cathrin = from plasma membrane between golgi and endoscopes
21
Q

How do we know what membrane is correct?

A

Rab-GTP

22
Q

What is Rab-GTP?

A

It is a GTP binding protein and GTPase found on the cytosolic surface of vesicles (60 types)
Rab proteins cycle between a membrane and the cytosol
GDP bound = INACTIVE (in cytosol)
GTP bound = ACTIVE (in vesicle membrane)

23
Q

How do we ensure specificity?

A

GEF found on vesicle membrane activates Rab - activates Rab-GDP in the cytosol to form Rab-GTP
Rab-GTP binds to Rab-effector protein on the target membrane = tethering of the vesicles to the target membrane

24
Q

What occurs during membrane fusion?

A
  • for vesicular cargo to reach the target organelle membranes must fuse = SNARE proteins
  • V-SNAREs (vesicle) and T-SNARES (target) are specific to one-another providing another layer of specificity
  • they wrap around each other to from stable trans-SNARE complex
  • once within 1.5nm phospholipids can flow between - fusion occurs
25
Q

What happens to Rab-GDP and Rab-GTP during membrane fusion?

A
  • A Rab-GAP (GTPase actvating protein) cause Rab to hydrolyse GTP to GDP
  • Rab-GDP is then released from the vesicle and bound to Rab-GTP dissociation inhibitor (GDI) to keep Rab inactive
26
Q

What is the Golgi apparatus?

A
  • stacks of flattened membrane enclosed compartments termed cisternae
  • Cis and Trans faces
  • CGN = recieves vesicles from the ER
27
Q

What are the function of the Golgi apparatus?

A

1- promote correct folding of proteins
2- prevent unwanted aggregation
3- acts as signals for sorting and targeting to the correct pathway

28
Q

What are lysosomes and their function?

A
  • lysosomes are degradative organelles - digesting unwanted material
  • contains lotrs of enzymes to break down macromolecules
  • ony active at pH 4.5 - 5 this is important for cell protection
  • pH is maintained via vascular ATPase
  • must be delivered via the golgi in clathrin coated vesicles
  • mannose-6 phosphate tagged
29
Q

What is endocytosis?

A

after an endocytic vesicle fuses with an early endosome the ingest material can be either:
1- DEGRADED = moves towards centre of cell and membrane composition alters. No longer recycles. Starts degradation
2 - RECYCLED = vesicles containing material for recycling to the plasma membrane bud off and fuse with a recycling endosome

Biochemistry (5 decks)

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