GPCR Flashcards
Improperly folded proteins have the following effects (5)
Aggregation-prone Bind chaperones Non-functional (improper interactions or non-productive interactions) Targeted to degradation Resource Drain : energy and material
What is the ER signal sequence?
Protein sequence on the N-terminal that allows ER import
How does the GPCR have so many loops inside the membrane?
multiple stop and start sequences
General role of molecular chaperones
Assist proteins during their maturation
What do the molecular chaperones bind to on the protein so assure proper maturation?
Hydrophobic segments that could potentially bind to these same segments on other proteins
If hydrophobic segments from different proteins bind, what does this cause?
aggregation
3 functions of chaperones
General folding helpers
Formation of disulfide bridges
Enzymes of the quality control cycle
Example of a chaperone that helps folding, what does it do?
BiP : prevents premature and incorrect folding of segments that arrive in ER lumen
Example of chaperone that helps the formation of disulfide bridges
Protein Disulfide Isomerase (PDI)
Two enzymes of the quality control cycle? what do they do?
Calnexin (membrane)
Calreticulin (soluble)
They bind to glucosylated oligosaccharides of incompletely folded proteins and prevent aggregation until PDI arrives
What is glycosylation?
Addition of oligosaccharides
The addition of oligosaccharides (glycosylation) serves as what?
The presence of sugars is used to monitor the _____ state of a protein
a tag to mark the state of protein folding
folded
What enzyme does glycosylation?
Oligosaccharyl transferase
Quality control cycle:
What happens after addition of sugars? This leaves what as a substrate? What chaperone acts on that substrate?
Trimming in the ER (removal of 2-3 outer chain glucose)
Leaves one glucose
Calnexin acts on glucose
Quality control cycle:
After action of calnexin on the single glucose substrate, which enzyme comes into play and what is their role?
Glucosidase
Removes remaining glucose and allows the protein to continue folding
Quality control cycle:
What happens if the protein is incompletely folded after already going through a quality control cycle?
A single terminal glucose is readded by glucosyl transferase and a calnexin is regenerated. This is repeated until protein is properly folded.
4 modifications to create a functional receptor
Glycosylation
Palmitoylation
Disulfide bridges
Dimerization
Which chaperone is important for dimerization of GPCR?
Dimer-probing chaperone
Where does dimerization of GPCR happen?
Membrane of ER
What happens to unfolded and monomeric GPCRs?
Degraded by ER-associated degradation pathway
After GPCR dimerization at ER, GPCRs are matured in the ____
Golgi
After maturation of GPCR at Golgi, GPCR homodimers go to the ____________ where they interact with ______
plasma membrane
heterotrimeric G protein
5 GPCR dimerization functions
Ontogeny Ligand-promoted regulation Pharmacological diversity Signal transduction Internalization
3 ER retention motifs?
KDEL
KKXX
RXR
which proteins have KDEL motif?
ER luminal chaperone proteins (BiP + PDI)
with proteins have KKXX motif?
type I integral membrane proteins
Role of KDEL and KKXX motifs?
Recycling proteins from Golgi back to ER
RXR motif found where?
several GPCRs
Role of RXR motif?
Precludes exit of proteins from the ER
Two proteins involved in trafficking
SNARE proteins
Rab GTPases
Role of SNAREs?
Recognition and catalyze fusion of transport vesicles with target membrane
Role of Rab GTPases?
Regulate docking and tethering of the transport vesicle to the target membrane.
Rabs in GDP-bound state are ____ and at ______
inactive
cytosol
Rabs in GTP-bound state are _____ and at _____
active
membrane of an organelle or transport vesicle
True or False
Each different Rab protein controls a different transport pathway between the ER and plasma membrane
True, different proteins control different pathways
True or False :
Each Rab protein has a specific SNARE protein that will target them
True
GPCRs can ______ or bind to ______
dimerize
G protein
GPCR’s final destination is the ________
plasma membrane
3 steps in signal transduction regulation
Desensitization
Sequestration
Recycling/Downregulation
What is desensitization and why does it happen?
Removal of receptor so it cannot accept more hormone/signal
What happens in the desensitization step of signal transduction regulation?
Phosphorylation of receptor by GRK2
Arrestin recruitment
Clathrin recruitment and coats plasma membrane
Dynamin surrounds neck of vesicle and pinches it off
Sequestration : two classes of GPCRs?
Class A
Class B
Sequestration : what does the Class A GPCR do?
Removal of arrestin and hormone so the receptor can be reused
Sequestration : what does the Class B GPCR do?
Slow recycling?
Degradation
Recycling/Downregulation : what is the mechanism?
Dephosphorylation of receptor by PP2A
Removal from it’s ligand
True or False :
Receptor localization is static
False
The vesicle that gets pinched off to eventually get recycled is called the ________
early endosome
Clathrin-coated pits internalization:
What are to components involved in the coat nucleation and assembly?
AP-2 AP180 Clathrin Synaptotagmin PIP2 Cargo
Clathrin-coated pits internalization:
What are the components involved in the coated pit maturation?
Dynamin
Endophilin
Amphiphysin
Actin
Clathrin-coated pits internalization:
What are the components involved in Fission of the vesicle?
Dynamin
Endophilin
Clathrin-coated pits internalization:
What are the componentsa involved in uncoating?
Hsc70
Auxilin
Synaptojanin
Stoned B
Do early endosomes have lower or higher pH and why?
Lower so it can release the receptor and ligand
What are the markers in the early endosome?
EEA-1 proteins
rab5-GDP
Role of recycling endosomes?
Return the receptor to the cell surface by binding to the plasma membrane
Marker for recycling endosome?
Rab4-GDP
Late endosomes : low or high pH
Low
Markers for late endosomes
rab7-GDP
LBPA
MPR+
Can late endosomes digest all materials? If not, what do they do? What is the marker for this?
No
Fusion to lysosome
LAMPs
Enzymes in the lysosome
Acid hydrolases Lipases Carbohydrases Proteases Nucleases Phosphoric acid monoesters
Markers for the lysosome
LAMP+
Acid hydrolases
MPR negative
Why would a receptor have to be degraded rather than recycled? (3)
If it requires cleavage during activation
If it is bound to irreversible ligands
Proteins that are not recycled back to plasma membrane, coming from the endocytic pathway
If there is accumulation of proteins in the cytosol, what does this affect?
Transport to ER
If there is accumulation of protein in the rough ER, what does this affect?
Budding of vesicles from rough ER
If there is accumulation of protein in the ER-to-Golgi transport vesicles, what does this affect?
Fusion of transport vesicles with Golgi
If there is accumulation of protein in the Golgi, what does this affect?
Transport from Golgi to secretory vesicles
If there is accumulation of protein in the secretory vesicles, what does this affect?
Transport from the secretory vesicles to the cell surface
What are the two fates of misfolded proteins?
Ubiquitination and degradation by proteasome
Molecular chaperones remodel the protein
Which diseases cause misfolding? (4)
CF
Marfan Syndrome
Nephrogenic Diabetes Insipidus
Alpha-1-Antitrypsin Deficiency
Which diseases cause retention in the ER?
CF
Nephrogenic Diabetes Insipidus
Alpha-1-Antitrypsin Deficiency
Which disease cause aggregation in the brain after protein release?
Creutzfeldt-Jakob Disease
Alzheimer’s
Phamacological chaperones role?
Stabilize protein conformation so it can redirected to the plasma membrane rather then to the proteasome/lysosome for degradation
Lack of proper folding results in what? (3)
Aggregation
Intracellular retention of proteins that shouldn’t be there
Destruction of proteins that should be expressed
Mistrafficking provokes ______ and ______ of essential proteins
Intracellular retention
Degradation
