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27 Cards in this Set
- Front
- Back
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secretory pathway - protein trafficking |
synthesized in ER by ribosomes processed in golgi secreted in vesicles (constitutive or regulated) |
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pulse chase |
technique to study trafficking cells are incubated with "hot" amino acid (pulse) "hot" amino acid is replaced with "cold" observe just the proteins that were made during pulse and where these proteins go in the cell ER->golgi->apical vesicles |
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green flourscent protein (GFP) |
purple loops change their conformation when they absorb energy, when energy is released and they go back to normal conformation energy is released as light add gfp dna sequence to the end of dna sequence of interest protein takes gfp wherever it goes |
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gfp and temperature sensitive mutations |
yeast that has a real mutation but at some temps, even though protein is mutant, it still functions take the cell to a higher temperature, protein doesn't function as well **AA sequences that are altered in mutant are required for the protein to move to the golgi |
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cell free systems |
used to study trafficking microsomes separated by density antibodies generated against proteins antibodies used to co-stain cells as endomembrane markers of protein trafficking determine co-localization |
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vesicle budding |
'bud' from the ER 'fuse' to the golgi |
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rough ER |
ribosomes many flattened membranes continuous with outer membrane of nucleus synthesis of exported proteins cells of salivary glands, intestine = polarized "tear drop" - secretes only on one side |
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other functions of RER |
initial steps of addition of carbohydrates to form glycoproteins folding of polypeptides recognition and removal of misfolded proteins assembly of multimeric protein complexes |
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smooth er |
no ribosomes tubular membrane system activities within depend on cell type cytochrome p450 (superfam) puts O on hydrocarbon to make it hydrophilic enough to be excreted |
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signal hypothesis and site of protein synthesis |
to be secreted have signal sequence of AA at their N-terminal, says "take me to ER" SS emerges from ribosome, bound by SRP SRP moves ribo to RER, bound by SRP receptor and TM channel (translocon) where protein is let into Er SS cleaves off, chaperones help protein to fold correctly
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what happens in the lumen of the ER |
protein processing begins- signal peptide cleaved (signal peptidase) protein glycoslyation begins (oligosaccryltransferase) chaperones assist in folding intramolecular disulfide bonds are formed (PDI) - btw 2 Cys |
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chaperone proteins |
folding machinery = proteins which assist new synthesized proteins in folding recognition = proteins recognized misfolded, if can't be resolved --> destroyed by proteosome |
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ERAD - recognition machinery |
employed for 1 or few bad proteins, depends on glycosylation having already happened put on sugar, 2 enzymes take it off 1 glucose bound by calnexin now other proteins check for exposed AA last Glc removed, if AA exposed, Glc added back, back to calnexin, checked again, if folded correctly, sent to er processed through golgi if not folded correctly ->proteosome for degradation |
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unfolded protein response - recognition machinery |
when misfolded back up in ER depends on activation of misfolded protein sensors, kept apart/inactive by BiP
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UPR - 2 types activated sensors |
dimers - phosphorylate eachother, stops protein translation
cleavage of cytosolic portion of sensor which translocates to nucleus where genes are transcribed to relieve ER of stress |
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protein glycosylation in the RER |
branched oligosaccarides are synthesized, transferred to Asp residue of protein by oligosaccharyl-transferase synthesis begins on lipid carrier - dolichol phosphate |
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ERGIC anterograde retrograde |
how proteins get from ER to golgi vesicles bud from ER and fuse to form larger ERGIC on the way some proteins keep going and some go back to the ER travel along microtubules anterograde = ER to golgi retrograde = golgi to ER traffic |
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anatomy of golgi |
2 networks, 3 levels of cisternae cis = closer to ER trans = side of release of vesicles network levels sort proteins CGN = desination TGN = right vesicle |
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membrane stacks of golgi supported by... |
cytoskeletal proteins: actin, spectrin, and ankryin |
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complex glycosylation reactions in golgi |
mannose removed and replaced by other sugars order of sugar attachment achieved by spatial arrangement of transferases AA sequence around sugar attachment tells the golgi to add particular sugars proteoglycans completely assembled in golgi |
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COPII coated vesicles |
anterograde traffic proteins that bind to TM proteins and lipids bearing an ER "signal peptide"
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COPI coated vesiles |
retrograde also "retrieve" ER proteins and enzymes that have been transported to golgi use GTP-binding coat protein (Arf1) peptide signal KDEL indicates ER resident protein and binds to KDEL receptor kdel receptor has peptide signal KKXX binds to COPI protein thus, all KDEL proteins are returned to ER via COPI |
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transport and fusing of vesicles |
use microtubules as tracks "tethered" to target compartment (Rab proteins position vesicle over SNAREs) vSNAREs (vesicle) bind to tSNAREs (target) for docking signal brings target and vesicle together uncoating process in order to fuse |
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receptor mediated endocytosis |
clathrin and ap2 induce curvature transport vesicle goes to endosome for sorting |
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early endosome |
sort housekeeping from signaling receptors and ligands |
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late endosome |
closer to TGN contain the cargo recycling compartments - some proteins go back to PM |
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bulk phase endocytosis |
pinocytosis appears non-specific any molecules near clathrin coated pit are internalized |
