Flashcards in Co-Translational Translocation (continued) Deck (42):
What is the function of N terminus signal sequence in translocation? What else is it known as?
The N terminus signal sequence is also known as the start transfer sequence. This sequence 'tells' a protein to go into the protein translocator.
If there is no hydrophobic sequence long enough to be a transmembrane in the protein (other then the signal sequence/start transfer signal), then what will happen to the signal sequence? What about the rest of the protein?
The signal sequence will be cleaved by a transmembrane peptidase. The signal sequence will be laterally released into the membrane. The rest of the protein will be released into the lumen of the ER.
If we have a protein with a signal sequence/start transfer signal at the N-terminus, and a Non-Polar sequence of sufficient size somewhere in the middle,
What is another name for the Non-polar region in this case? What will happen to the signal sequence, What will happen to the protein?
The Stop-Transfer sequence.
Once the stop-transfer sequence enters the transmembrane region, the start transfer sequence will be cut by a peptidase, removed into membrane laterally.
The rest of the protein will also be moved laterally into the membrane.
What is the lateral gating mechanism?
The release of a protein into the plasma membrane via an alteration of the shape of the protein translocator due to the entry of the hydrophobic stop transfer sequence. The end result is the opening of a gate in the translocator protein and the release of the protein being translocated into the membrane through this gate.
Is it possible to have an internal start transfer sequence?
What is the fate of this internal sequence?
It will be sent into the membrane. No cutting with peptidase.
Is the lumen of the ER more positive or negative then the cytosol?
The Lumen of the ER is more positive (likely due to Ca2+) then the cytoplasm.
What dictates whether a protein with an internal transfer sequence has its N or C terminus in the lumen of the ER?
The transfer sequence will be flanked by charged amino acids. If the C terminus contains a - charge (the N will have a +) and the C terminus will be inserted into the more positive lumen of the ER.
How are two transmembrane domains inserted into the membrane?
The protein will have an internal start transfer sequence (it may or may not have an N terminus start transfer sequence as well). This will stay in the membrane, and a subsequent stop transfer sequence will cause opening of lateral gate and release of the protein, with both transfer sequences in the membrane.
If there are two transmembrane sequences which one dictates whether the N terminus and C terminus resides in the Cytosol or ER lumen.
It is the flanking charged amino acids on the membranes start sequence. which dictates placement. This will be true regardless of the number of transmembrane sequences in the protein.
How are multiple transmembrane domains inserted into the membrane?
The Protein translocator treats them like alternating stop and start transfer sequences. The first start sequences flanking amino acids charge decides orientation. Afterwards each start and stop transfer sequence is woven into the membrane.
what does the pre-targeting complex do?
It recognizes new proteins with a special C-terminus hydrophobic region, and helps it bind to get-3 ATPase.
What does get-3 ATPase do? Where is it located?
Once it has been transferred a protein by a pre-targeting complex. It will find the get1-get2 complex, which resides in the ER membrane. And give it the protein. Get3 atpase will then hydrolyze ATP and disassociate... apparently it was a dimer held together by ATP. Get3 resides in the cytosol.
What does the get1-get-2 complex do?
It deposits the hydrophobic C terminus of a protein into the ER membrane, so that the protein is facing into the cytosol. This complex is membrane bound.
How many types of protein glycosylation are there?
Two, N-type and O-type.
What is the consensus sequence for N-type glycosylation?
N(asn) - x - S/T (Ser/Thr)
What does the glycosylation use as its attaching point?
It attaches to the N in the amide side group of Asn.
Describe the structure of the glycosylation, from most proximal to the N in the amide to most distal.
N-acetylglycosamine = N
Mannose = M
Glucose = G
N } N-acetylglucosamine.
M, M }
M, M, M } Mannose
M, M, M }
G } Glucose
Which enzyme recognizes the N-X-S/T consensus sequence? Where does recognition occur?
In the Lumen of the ER.
What is dolichol?
It is a lipid anchor glycosylation. (so it will become a glycolipid)
How is glycolipids synthesized (assuming a dolichol membrane attachment).
Dilochol (d) starts with its glycosidic side facing the cytosol.
Phosphorylated by CTP
Then phosphorylated by UTP, and two N-acetylglucosamines are added.
(GTP --> GDP adding manose.) 5X
Flips in membrane.
(Mannose donor gives Mannose. ) 4x
(Glucose donor gives glucose) 3x
How does glucose trimming act as a control for protein folding.
A protein which is glycosylated has its 2 of 3 glucoses cut off.
calnexin removes the final glucose, proteins tries to fold if it succeeds it leaves.
If it doesn't succeed one glucose is added back on by glucosyl transferase. And the cycle begins again.
What does calnexin do.
It removes the final glucose, then the protein has another chance to fold.
What does glucosyl transferase do.
It transfers a glucose onto something.
How does glycosyltransferase know whether or not it should add a glucose?
It does not 'recognize'/bind with affinity to the folded protein.
What happens to proteins that have been mis-folded irreparably?
They are guided by a chaperone to a protein translocator/ubiquiten ligase. Which transports it out, and adds ubiquitin to mark it. N-glycanase cuts off its oligosaccharide, and a proteasome eats it.
What is a lectin?
A class of proteins which bind to sugars, and cause them to clump.
How does the cell regulate genes related to protein folding capacity?
There are sensors in the ER membrane which bind misfolded proteins and signal to increase transcription of genes which regulate proteins involved in protein folding.
What is IRE1? What does it do?
IRE one detects unfolded proteins in the ER, allows introns to be cut out of transcription regulators that help to deal with high levels of unfolded proteins.
Proteins with a GPI (glycosylphosphatidylinositol) anchor will be attached to an inositol.
Where are proteins with a GPI anchor found at after transport?
The surface of the extracellular membrane.
What happens when GPI-anchored proteins end up in the lipid rafts?
They make signaling hotspots.
Trypanosoma defense mechanism involves interfering with what?
produces a phospholipase which cuts the GPI anchor. In red blood cells. It is parasite that causes sleeping sickness and chagas disease.
where does synthesis of phospholipids occur?
cytosolic layer of the ER membrane.
where does synthesis of phospholipids occur?
CoA is added to two fatty acids, which are both then attached to glycerol 3 phosphate.
Phosphate is hydrolyzed, and then a head group can be attached which contains its own phosphate.
Where is scramblase found?
It is a transmembrane protein in the ER membrane.
What does scramblase do?
It non selectively (non specific) flips phospholipids across the ER membrane. Keeping the cytosolic monolayer from growing too much. It equalizes the composition of the monolayers and is constitutively-active.
What does flippase in the PM do?
It selectively flips phosphatidylethanolamine and phosphatidylserine to the cytosolic layer of the cell.
Is plasma membrane flippase constitutively active?
No. It is deactivated to signal apoptosis.
serine + 1 fatty acid =
serine + 2 fatty acids =