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Cell Biology > Membrane Fusion > Flashcards

Membrane Fusion Flashcards

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1
Q

what is the concept behind membrane fusion?

A
  • cells are compartmentalized but need movement
  • requires membrane fusion
  • displacement of water from the hydrophillic surface of membrane is energetically unfavourable
  • requires SNARE protein
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2
Q

why do membranes repel each other?

A

phosphates have a negative charge so phospholipids repel each other

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3
Q

what is membrane fusion?

A

the process where two separate lipid bilayers merge to become one
- need to be brought together closer than 1.5nm

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4
Q

what do membranes contains?

A

lipids and proteins

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5
Q

describe the vesicle budding

A
  • contains part of the lumenal contents of the donor membrane
  • contains membrane proteins and lipids from donor membrane
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6
Q

how does the vesicle fuse?

A
  • contents mixes with the lumen of the accepting membrane or is released extracellularly
  • proteins and lipids become part of the accepting membrane
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7
Q

what are the 3 steps of fusion?

A
  1. tethering - bridge between two membranes
  2. docking/the formation of a trans-SNARE complex
    - fusion
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8
Q

what are the different stages of fusion?

A
  • fusion initiation
  • hemi fusion
  • fusion pore opening
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9
Q

what are MTCs?

A
  • multi subunit tethering complexes that provide a protein bridge
  • tether one membrane to another
  • bind a verity proteins on opposing membranes, this provides some specifity
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10
Q

what i the role of SNARE proteins?

A
  • proteins that bring the membranes into close proximity

- allows membranes to overcome the repulsive forces

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11
Q

what is the structure of SNARE proteins?

A
  • most are transmembrane
  • contain a SNARE domain
  • when it comes into contact another SNARE protein they form an alpha helical coil
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12
Q

what is a SNARE complex?

A
  • a SNARE complex consists of 4 helical coils
  • either 4 individual or a SNAP proteins and 2 others
  • SNARE proteins are unstructured until they form a SNARE complex
  • 4 helices are usually one helix from one membrane and 3 from another
  • one of the strongest complexes
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13
Q

what is zippering?

A

zippering of the SNAREs overcomes the electrostatic repulsion of the phospholipids

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14
Q

what is post-tethering?

A

post tethering the SNAREs on opposite membranes starts to interact and then coil up forming the SNARE commplexing
- if you coil up a protein it becomes shorter, bringin the membranes into close proximity and overcoming the membrane repulsive forces

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15
Q

what is the centre of the SNARE complex?

A
  • 4 amino acids, one from each helix pointing inwards
  • either glutamine (Q) or arginine
  • always 3Q’s and 1R
  • Q SNAREs, Qa, Qb, Qc
  • lots of hydrogen bonding and salt bridges
  • hydrophobic central core
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16
Q

how is the SNARE complex specific?

A
  • you need the right combinations of SNARE proteins for membrane fusion
  • Qa,Qb,Qc and R SNARE are required for every membrane fusion event
17
Q

how are SNAREs the minimal machinery for membrane fusion?

A
  • experiment show that if you can make the SNAREs type I membrane proteins can get cells to fuse with each other
18
Q

describe the process of SNARE proteins fusing (the experiment)

A
  • SNARE domains need to be in the cytoplasm for membrane fusion
  • N terminus in cytoplasm and C terminus in the lumen are type II membrane proteins
  • converted them into type II membrane proteins
  • flipped v SNARE (made the cytoplasm red)
  • flipped t SNARE (made the nucleus blue)
  • cells then fused - had multinucleated cells
19
Q

why is the SNARE complex good for fusion?

A

once formed it is extremely stable

20
Q

how are you abale to resuse the SNARE proteins/

A
  • need to break the stable complex

- with a ALPHA-SNAP and NSF

21
Q

what is NSF?

A
  • hexamer
  • ATPase, hydrolyses ATP
  • Sensitive to NEM
22
Q

what is alpha SNAP?

A
  • required for NSF bidning to membranes

- binds and activates NSF ATPase activity

23
Q

how do NSF and alpha SNAP work?

A

form a complex and with hydrolysis of ATP break open the SNARE complex
- SNARES bind to SNAP, they are SNAP receptors

24
Q

what happens if you inhibit NSF?

A

you would still get one round of fusion

25
Q

how are SNAREs regulated?

A
  • sec1/Munc18 (SM proteins) and can regulate SNARE assembly
  • when bound to sec1 it is in a closed conformation the SNARE protein would not be available to form a SNARE complex
  • can clamp SNARE proteins
26
Q

what are Rab proteins?

A
  • prime membranes for fusion
  • not fully understood
  • family of 60 genes in humans
  • interact reversibly with membranes and effectors in a GTP dependent manner
27
Q

what is bound to inactive Rab proteins?

A

GDP

28
Q

what is bound to active Rab proteins?

A

GTP

29
Q

what do Rab proteins do?

A

it has low intrinsic GTPase activity, need GAP proteins (activating proteins)

30
Q

what do you need to exchange GDP to GTP?

A

GEF proteins - exchange factor

31
Q

what do Ras do?

A

makes membranes permissive

32
Q

what do all fusion events have?

A

4 helix SNARE complex, an SM protein and a Rab protein

Cell Biology (10 decks)

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