Biochem 3 - Liu, Yokomori, Donovan Flashcards
What proteins are the major components of Clathrin coated vesicles and what are their functions?
Clathrin (provide the physical force of budding and vesicle formation and make a network of pentagons and hexagons on clathrin cage - triskelion) and adaptin (bridges clathrin to transmembrane receptor)
Role of dynamin
GTPase that helps pinch off clathrin coated vesicle
Function of Sar1 and ARF
Sar1 for Cop II and ARF for Cop I and Clathrin. Coat-recruitment GTPases are usually in the cytosol in an inactive, GDP-bound state. Sar1 is normally a cytoplasmic protein that has an attached myristic acid buried inside of the protein. In the cytosol it is found in the GDP-bound form - the myristic acid is probably sequestered in the protein somehow. When a COPII- coated vesicle is to bud from the ER membrane, a specific GEF embedded in the ER membrane binds to cytosolic Sar1, causing the Sar1 to release its GDP and bind GTP in its place, and inducing it to bind to the donor membrane by insertion of its myristic acid into the lipid membrane. Sar1-GTP bound to the donor membrane triggers the coatomer proteins to bind, cluster sar1 and form a bud. Coatomers will bind to its target membrane only after the membrane has been primed by Sar1. Other GEFs and coat-recruitment GTPases operate in a similar way on other membranes.
SNARE hypothesis
There is a membrane protein on the vesicle called v-SNARE and there is a membrane protein on the target membrane called t-SNARE. The fitness between a v-SNARE with a complementary t-SNARE determines the specificity of docking. The fitting might be monitored by a member of the monomeric GTPase family called Rab protein. In this view Rab proteins become attached to the surface of coated vesicle in the donor membrane. When vesicle encounters the correct target membrane, the binding of v- SNARE to t-SNARE causes the vesicle to remain bound for long enough to allow the Rab protein to hydrolyze its bound GTP, which locks the vesicle onto the target membrane. These together with NSF induce the two membranes to fuse.
How are SNAREs pried apart after fusion?
A cytosolic protein NSF + ATP is required for prie apart interacting SNAREs before they can function again.
What entry mechanism have viruses adopted that’s similar to vesicular transport?
Fusion mechanism: The interaction of a chemokine receptor on target cell with the viral GP120 protein release the previously buried hydrophobic fusion peptide to insert into the plasma membrane of the target cell. The fusion protein, now anchored in the two opposing membranes, rearrange its structure, and the energy released from this rearrangement pulls the two membrane together, like a mouse trap.
Transport from the ER to the Golgi is mediated by_____
Vesicular Tubular Cluster via COP II vesicles
How can you have all this membrane transport from the ER to the Golgi to the plasma membrane without depleting the whole ER and Golgi?
The answer to these questions is that there is retrieval or retrograde transport. As soon as vesicular tubular clusters form, they begin budding of as COPI coated vesicles. They carry the ER membrane and ER resident proteins that have escaped back to ER.
Retrieval of escaped ER proteins depends on _____
ER retrieval signal. Resident ER membrane proteins have signal sequences that bind to COPI and are thus packaged into COPI-coated vesicles for retrograde delivery to the ER. Resident ER proteins have a different short sequence Lys-Asp-Glu-Leu (KDEL). If this sequence was removed, the proteins were not retained, and if this sequence was placed on the C-terminus of normally secreted proteins they remain in the ER. The KDEL sequence binds to specialized receptors, such as the KDEL receptor, a transmembrane protein that binds to KDEL sequences and packages any proteins with KDEL sequence into COPI-coated retrograde transport vesicles. KDEL receptors recycle between the ER and the Golgi, retrieving ER resident proteins. Different affinities of the receptor binding to KDEL sequences in the ER and Golgi may be determined by their differences in pH. (Golgi more acidic)
Describe membrane recycling in Golgi
Membrane recycled back to Golgi as immature secretory vesicle matures. Membrane recycling is important for returning Golgi components to the Golgi apparatus, as well as for concentrating the contents of secretory vesicles. The vesicles that mediate this retrieval originate as clathrin-coated.
Describe docking system between synaptic vesicles and presynaptic membrane
So docking system between synaptic vesicles and presynaptic membrane is similar to vesicle transport between ER and the Golgi.
Protein sequences of synaptogamin and syntaxin are very similar to the t- SNARE and v-SNARE
Also associated with this complex is Rab3, a synaptic vesicle specific G- protein.
Where does phagocytosis occur?
Phagocytosis occurs only in specialized cells, such as macrophages and neutrophils.
2 types of pinocytosis. Where does pinocytosis not occur?
RBC; Fluid phase non-selective pinocytosis and Receptor-mediated selective pinocytosis (occurs in coated pits, most often clathrin)
What happens when the LDL receptor does not accumulate in coated pits?
familial hypercholesterolaemia; Class of mutations in the c-terminal cytoplasmic domain - This signal sequence, Phe-Arg-X-Tyr, on the C-terminus of LDL receptor is for adaptin binding.
Adaptins serve as the molecular link between clathrin-coats and specific transmembrane receptors. Generally, the clathrin-cages are the same in each case; but the adaptins are different, and they mediate the capture of the different types of cargo receptors.
3 outcomes of endocytosis
- Receptor recycles: receptor preclustered in coated pits - recycles even in the absence of ligand. Examples:
- LDL receptor
- transferrin receptor. - Receptor degraded: receptor and ligand are both degraded. Example: The Epidermal Growth Factor Receptor (EGF Receptor) (and other growth factor receptors like insulin receptor)
- Transcytosis: receptor on the surface of polarized epithelial cells transfer specific macromolecules from one extracellular space to another. Example: Maternal IgG Receptor. New born mammals absorb maternal IgGs from their mother’s milk. This is done by receptor mediated endocytosis in the intestinal epithelial cells. The gut is at about pH 6.0 and IgG binds to the receptor. The extracellular fluid in circulation is at pH 7 where the IgG dissociates from the receptor.
