MCB Lecture 32 Vesicular Transport Flashcards
What are the features of the Stop Transfer Sequence?
- Hydrophobic
2. Resistant to peptidase, isn’t cleaved by peptidase
Describe how transmembrane proteins become embedded in the membrane
- START transfer sequence present on N terminus, as well as STOP transfer sequence present further up
- Start transfer sequence binds to the translocator protein bound in the ER membrane
- Protein is fed through until it reaches the STOP sequence
- Signal peptidase cleaves the START transfer sequence from the rest of the protein.
- Translocator dissociates, leaving the protein embedded at the STOP sequence
Describe the orientation of transmembrane proteins and how this can be changed
Normally, N terminus is inside, C terminus is outside
To invert the orientation, the START transfer sequence is present further up from the N terminus.
The protein binds to the translocator here, with the N terminus still outside the cell
The protein is fed through, until the C terminus is inside the cell
Describe how multipass proteins become embedded in the membrane
There are multiple START and STOP domains
Why does glycosylation occur?
This tracks the folding of the protein
Describe the process of glycosylation
- Oligosaccharyl transferase moves sugars from Dolichol to a Asn residue on the protein as soon asit enters the ER
- N-acetyl glucosamine, mannose, and 3 glucoses are added
Describe the sugars on an unfolded protein
All three glucose molecules are attached
Describe the sugars on an incompletely folded protein
One glucose remains attached
Describe what happens when a protein does not fold properly
A glucose molecule will be reattached by glucosyl transferase, and then will bind with calnexin again
What is calnexin?
It is a protein bound in the ER membrane.
It binds to the glucose on a protein and helps it fold
The glucose is then cleaved
Describe the process of protein degredation
When a protein misfolds:
- A chaperone bindso
- It is transported out of the ER back into the cytosol
- Ubiquitin binds to it
- The ubiquitin tagged protein is sent to the proteosome, which degrades it
What is the name of the protein that cleaves the signal sequence
Signal peptidase
What is dolichol?
It is a lipid in the membrane of the ER with the oligosaccharides that end up of the protein to be folded
What is oligosaccharyl transferase?
It transfers the oligosaccharides group from dolichol to the protein
What are the three different pathways that use vesicular transport?
- Endocytic
- Biosynthetic (secretory)
- Recycling/retrieval
Describe the biosynthetic pathway
Molecules are translated and folded in the ER.
Vesicles bud of and are delivered to the Golgi
The proteins are packaged and altered in the Golgi
From the Golgi, vesicles bud off and are exocytosed
What are the three vesicle coat proteins?
Clathrin
COP1
COPII
Describe the endocytic pathway. How do vesicles form?
Endocrine pathway is the endocytosis of stuff from outside the cell. These phagocytotic vesicles become early endosome a, then late endosomes, then lysosomes for degradation of the contents.
- Cargo receptors are bound in the membrane
- Adaptins bind to the triskelion of the clathrin molecule and the cargo receptor
- Clathrin makes the membrane curve out, and concentrates cargo receptors in the clathrin coated pit
- Dynamin destabilises the narrowing, so that the vesicle blebs off. (GTP dependent process)
What is clathrin?
Clathrin is a vesicle coat protein that forms vesicles in the endocytic pathway
How are vesicles targeted to specific organelles?
There are proteins bound on the outside of the vesicle that moderate targeting.
- Rab and v-SNARE are bound to the external side of the vesicle
- Tethering molecule on the cell membrane interacts with Rab, bring the vesicle into close proximity
- v-SNARE and t-SNARE on the target cell membrane interact, and the vesicle fuses
What is Rab-GTP?
It is a protein on the outside of a vesicle that is specific to certain organelles, leading to the targeting of organelles to specific organelles
What is v-SNARE?
This is the vesicle snare molecule that is involved with fusion of the vesicle to the organelle
What is t-SNARE?
It is the protein on the cell membrane of a vesicle that is involved with fusion of the vesicle to the cell organelle membrane
How is specificity of vesicle targeting maintained?
There are many different Rab proteins that are specific to different organelles
Describe vesicular transport from the ER to the Golgi
Vesicles form with COPII
These vesicles are targeted to the Golgi
The Golgi apparatus is a series of …
Ordered compartments
Each compartment in the Golgi has …
Specific enzymes involved with processing of enzymes
Describe processing in the Golgi
In each compartment, different additions are made to the proteins, due to the specific enzymes present in the various compartments
By the time a protein reaches the trans face of the Golgi, it has … attached
Negatively charged sugars attached
What are the two hypotheses for transport through the Golgi apparatus?
- Vesicular transport
- Cisternal maturation: vesicles from the ER fuse to form a cisterna. This cisterna moves along and gets incorporated into the Golgi complex
Where does glycosylation occur?
In the membrane of the rER
When is COP I used?
Formation of vesicles moving between the cisternae of the Golgi and formation of vesicles at the trans face for secretion
What regulates the Uncoating of vesicles once they have formed?
Cytosolic proteins
What happens to proteins with the KDEL sequence?
They are recycled from the Golgi to the ER in COP I coated vesicles
What is the main thing that happens to proteins in the Golgi?
Addition and Modification of oligosaccharides
To which residue are the dolichol oligosaccharides added during glycosylation in the ER?
Asparagine
