SNARE regulatory proteins Flashcards
what are SM proteins?
proteins whihcbintercat with syntaxin and SNARE pins
-are essential for membrane fusion
what is the shape of SM protein?
horseshoe shape
- 4 isoforms
what does Munc 18 do?
It is the binding partner for syntaxin (closed)
- bind to closed syntaxin to stop it from forming an unproductive SNARE complex
- negative regulator of secretion
How many binding sites are thought to be on munc18?
2.
- one binds to closed syntaxin and keeps closed
- one binds to open syntaxin - form SNARE
Is munc 18 required for vesicle fusion and how do we know this?
- yes
- when munc 18 is not present in artificial vesicles - rate of fusion is slow
- when munc18 pre-incubated then warmed up in artifical vesicles - fusion occurs at a quicker rate
- munc 18 is facilitating fusion
how can artificial vesicles be used to see if fusion is occuring?
- one vesicle contains flourescent material (cargo) and when the vesicles fuse this flouresence is quenched
(flouresence in quenching (t-vesicle) measured at % of maximum )
what happens in mouse chromaffin cells when munc-18 is knocked out?
- is no fusion of vesicles
- when munc-18 is re-expressed by a viral vector fusion is fully restored
- total number of vesicles normal - but proportion of docked vesicles is reduced
what does overexpression of SNAP-25 do in munc-18 KO chromaffin mouse cells?
-SNAP-25 rescues docking of vesicles but does not rescue fusion - shows that munc-18 still required for fusion to occur
what is munc-13 required for?
fusion of glutamatergic synaptic vesicles
what happens in a munc-13 KO?
- no glutamatergic transmission
- readily releasable pool of vesicles depleted
- vesicles can dock but cannot fuse
- munc must have a function in between docking and fusion
what does munc 13 do to syntaxin?
Munc 13 can open “closed” syntaxin which promotes SNARE complex formation
what part of SNAREs form binding sites for sm proteins?
helixes facing outside of SNARE motif form binding sites for sm proteins
