Transport I Flashcards
What’s the function of the ER?
Synthesize lipids and proteins for distribution to many other organelles and PM
What’s the volume occupied by organelles in percentage of entire cell volume?
50%! 54% is cytosol
What is the fate of proteins headed to membrane bound compartments?
ER➡️Golgi ➡️PM
What is the signal hypothesis? Give experiments on necessity and sufficiency.
To sort to an organelle, protein needs signal sequence to specify protein destination.
Eg NLS: 8AA signal sequence.
Necessity: hook GFP to mutant and non mutant (NLS missing) p53 and see that GFP only in nucleus in wt.
Sufficient: hook NLS to GFP only. If goes into nucleus then NLS is sufficient for translocation into nucleus.
What does necessary mean?
The signal sequence is required for signaling of a protein
What is the size of molecules that can pass freely through nuclear pore complex?
- Molecules 40kD must have NLS (remain folded during transport)
- expo: use GFP and synthetically generate different sizes of protein. See what’s the biggest protein for free diffusion.
How is active transport through the nuclear pore complex regulated? Where is the active and inactive form of this regulator?
GtPase ran! Small, monomeric cellular switch.
Gtp bound form is active. Hydrolysis of phosphate bind isn’t used for energy (unlike ATPase).
-ranGEF is bound to chromatin in nucleus (ranGTP is in nucleus)
-ranGAP is in cytoplasm
What is the role of ranGTP?
Nuclear import and export. RanGTP helps release import cargos by dissociating importin from the cargo. It also promotes cargo binding by binding to cargo and binding to exportin.
How do proteins enter the ER?
Co-translational translocation:
Proteins are trans located in unfolded form because these processes are simultaneous.
-SRP= signal recognition particle. Binds signal sequence on growing peptide that’s still attached to ribosome (slows translation)
- SRP binds receptor on ER and releases SRP
-peptide goes through translocation channel and into the ER lumen
How do soluble secretory proteins get through the membrane?
Translocation channel grabs signal sequence, signal peptide cleaved by signal peptidase, protein is soluble in ER and signal sequence remains in membrane.
How are single pass transmembrane proteins created?
Signal sequence at nh2 domain of peptide signals start transfer. Hydrophobic stop transfer sequence stops the flow of peptide into the ER lumen. Signal peptidase cleaves initial signal sequence. Hydrophobic stop sequence now passes through the membrane as the TM protein.
How are multi pass TM proteins made?
There’s an internal signal sequence that initiates transfer. And a second hydrophobic sequence that stops transfer. No cleavage. Both ends are in cytosol.
