Secretory Pathway and Protein Sorting Flashcards
What is the function of peroxisomes?
They are the site of oxidative reactions
What is the role of the endosome?
Recycling and sorting proteins. Also has a role in signalling
What experiments have been done to prove the cell is made up of compartments?
- Pulse chase experiments
- In Vitro Cell Free Assays
Outline the pulse chase experiments
- Cells are placed in a medium with 3H-leucine
- Cells are then placed in unlabelled medium
- Radioactive proteins are made and their position in the cell is tracked
- Cells are placed in radioactive medium only for enough time that one wave of proteins is labelled
Outline in vitro cell free assays
- Cells are broken upand fractionated by centrigation
- Organelles are then added back to the cytosol and ATP in order to see how proteins are trafficked
How can yeast mutants be used to help identify machinery of secretory pathways?
- Mutations introduced in yeast
- If the secretory pathway is interupted this would result in vesicles being stuck inside the cell at specific point
Give an example of a disease which is caused due to trafficking defects
Cystic Fibrosis
What causes cystic fibrosis?
Mutation in the CFTR chloride channel protein, causing protein to be trapped in the ER
If mRNA has no sorting signal where does the protein end up?
In the cytosol
What happens to mRNA with an organelle specific targeting sequence?
It is targeted and transported into the specific organelle
What type of sorting sequence leads to post-translational import?
Organelle specific
When does a protein have an ER signal sequence on the mRNA?
If it is a membrane protein or a protein to be secreted
What type of sorting sequence leads to co-translational import?
An ER signal sequence
What happens to proteins with an ER signal sequence?
Give an example of an experiment which proves the importance of signal sequences
Putting different signal sequences onto a cytosolic protein cause it to be expressed in different compartments in the cell
What is required from an ER signal sequence?
A string of hydrophobic amino acids (the actual sequence matters less, the hydrophobic nature is what is important)
What type of translocation is ER translocation?
Co-translational
What is the role of the ER?
- synthesis and folding of transmembrane and secreted proteins
- N-linked glycosylation
- GPI anchor attachment
- Lipid synthesis
- Calcium storage
What are the roles of the smooth ER?
- Lipid synthesis
- Where proteins exit the ER
What is the free ribosome cycle?
- There is a common pool of subunits in cytosol
- mRNA encoding a cytosolic protein remains free in cytosol
- Once the protein is built ribosomal subunits become free in cytosol again
What is the membrane bound ribosome cycle?
- Common pool of ribosomal subunits in cytosol
- if mRNA has an ER signal sequence then it binds an SRP which bind to SRP receptors, localising the ribosome to ER membrane
- Ribosomes translate mRNA and then return to cytosol
When a ribosome is brought to the ER membrane where is a ribosome localised to?
A translocation channel/translocon
Why is the translocon normally closed when not bound to the ribosome?
ER would otherwise be unable to hold Ca2+ in
What happens to the translocon when the ribosome binds?
- The translocon opens and the protein channel in the subunit lines up with it
- proteins synthesised in the MRNA are then directly cotranslated through the translocon into the ER lumen/membrane
