SFP: cellular basis of protein synthesis and secretion Flashcards
describe how proteins are destined for translation in membrane-bound ribosomes
they have a targeting sequence in their N-terminus that direct the ribosome to the rough ER
where does translation always begin
in the cytosol on free ribosomes
the outer layer of the ER is continuous with __. the lumen of the ER is continuous with ___.
the nuclear envelope; perinuclear space
list the basic steps involved in the secretory pathway
- mRNA goes to a translating free ribosome
- ribosome moves to the RER
- protein goes through the golgi complex
- protein enters a secretory vesicle
- protein is released from the cell via exocytosis
describe the function of the SRP?
a particle that binds to the signal sequence in the translating protein. it pauses translation of the protein and takes the whole protein-ribosome complex to the RER. the particle binds to the receptor, GTP converts to GDP, and the SRP is released.
what is the translocon?
the channel in the RER membrane where the ribosome binds during translation
once the ribosome and mRNA have been brought to the RER, where is the translating protein going?
into the lumen of the RER
describe the process of forming single pass transmembrane proteins in the RER
- Internal signal sequence (ISS) binds to SRP and causes a pause in translation
- SRP binds to receptor, SRP releases, ribosome binds to translocon, ISS is inserted into the translocon
- Translocon releases ISS laterally into the membrane and translation continues. the N or C terminus will poke into the cytosol. The AA sequence in the ISS determines how the growing protein is oriented.
describe the process of forming multipass transmembrane proteins in the RER
- 1st Internal signal sequence (ISS) binds to SRP and causes a pause in translation
- SRP binds to receptor, SRP releases, ribosome binds to translocon, 1st ISS is inserted into the translocon
- Translocon releases 1st ISS laterally into the membrane and translation continues
- 1st stop transfer sequence (STS) generated during translation pauses translation and releases STS into the membrane
- Translation resumes and 2nd ISS-SRP binds to the receptor, and a second loop in the membrane forms. This process of ISS and STS continues until the stop codon is reached and the protein is done.
name the various types of protein modification in the RER
N linked glycosylation, adding GPI anchors, protein folding
protein modification in RER: N linked glycosylation
oligosaccharide added to Asn in the lumen of RER. Occurs co-translationally. The oligosaccharide can be trimmed in the RER/golgi by various enzymes
protein modification in RER: GPI anchors
GPI anchors made in the luminal side of the RER membrane get added to membrane-spanning domains near the C terminus of specific proteins. The amino group off ethanolamine will be used to link the GPI anchor and the c terminus of the protein
where do GPI anchors in the membrane come from?
transported in vesicles from the RER that fuse with the plasma membrane
protein modification in RER: BiP and calnexin/calreticulin
involved in protein folding
- BiP: Hsp70 that binds to the protein as it enters the RER lumen
- Calnexin and calreticulin: recognize oligo chains of N linked glycoproteins
protein modification in RER: protein disulfide isomerase
enzyme in RER lumen that is important for correct protein folding by allowing for correct forming of disulfide bonds between cys residues; can break the bonds and let the correct ones reform
describe ER associated degradation (ERAD)
misfolded proteins are transported from RER to cytosol for degradation via ubiquitin-proteasome
describe the unfolded protein response
stress response from many unfolded proteins in the RER that activates a protein kinase pathway to stop translation
what is the tubular vesicular network?
the intermediate between the RER and cis golgi
what are the 4 main categories of protein processing in the golgi?
modifying N linked oligosaccharides, phosphorylating N linked oligosaccharides, O linked glycosylation, sulfation
protein processing in Golgi: modifying N linked oligosaccharides
in the cis/medial cisternae, the golgi can add GlcNac, galactose, and sialic acid and remove mannose. It does this for plasma membrane and secreted proteins
protein processing in Golgi: phosphorylation of N linked oligosaccharides
lysosomal enzymes get tagged with mannose-6-phosphate so they can be sorted into lysosomes. This happens in the cis Golgi network.
protein processing in Golgi: O linked glycosylation
sugar added to ser, thr, or tyr one at a time in the cis and medial cisternae
protein processing in Golgi: sulfation
sulfate added to tyr in the trans golgi network
which lipids can be made in the golgi, and where are they made?
sphingomyelin and glycolipids; luminal surface
what are the three secretory pathways in the golgi
constitutive (default pathway), regulated, and lysosome
secretory pathway: constitutive
- clathrin coated vesicle pinches off the golgi
- clathrin coat dissociates
- vesicle fuses to the plasma membrane
- material in the vesicle ends up on the cytoplasmic side of the plasma membrane
secretory pathway: regulated
- clathrin coated vesicle pinches off the golgi
- immature secretory granule forms (large vesicle)
- clathrin coat dissociates
- maturing secretory granule forms (bullseye appearance)
- mature secretory granule forms
- granule fuses and material releases through the membrane via exocytosis
secretory pathway: lysosome
- clathrin coated vesicle pinches off the membrane
- clathrin coat dissociates
- vesicle enters the late endosome
- lysosome is formed
what protein is needed to pinch off a vesicle? what is it?
dynamin; a GTPase
describe anterograde vs retrograde vesicular trafficking
-Anterograde: COPII protein coated RER vesicles bring stuff to cis golgi network or TVN
-Retrograde: COPI protein coated vesicles can move anywhere back from the trans golgi; can go all the way back to the RER
describe the basic process involved in sorting material into vesicles
- sorting sequence in protein binds to an adaptor protein
- adaptor protein found on inside of protein coat attaches the protein and the coat
- vesicle forms and moves to its destination
name the three common coats for protein-coated vesicles and the types of cargo they’re associated with
- COPI coat: luminal RER or RER membrane proteins
- COPII coat: membrane and secreted proteins
- clathrin: lysosomal proteins
briefly describe how exocytosis works
Target membrane has t SNAREs that bind to transport membrane v SNAREs. A RabGTP associated with the v SNARE hydrolyzes, allowing the two membranes to fuse.
