Main Ideas!- There are 3 basic mechs of protein transport!- gated transport (cytoplasm-nucleus)!- transmembrane transport (ER, mitochondria, peroxisomes)!- vesicular transport (ER-Golgi-lysosomes-extracellular space)!- Proteins are targeted across membranes via aa signal sequences!- With the biosynthetic-secretory pathway, soluble and membrane-bound proteins are shuttled via vesicle transport!- Protein glycosylation and oligosaccharide processing occur in the ER and Golgi!- Protein glycosylation is imp. for protein targeting to the lysosomes and the secretory pathway Learning Objectives 1) Know the essential road map of protein traffic - Cellular compartments: interior of a cellular compartment is basically the same all the way thru and the same as extracellular area, red= same interior compartment envi (mitochondria not part of compartmental comm. system ) 2) Understand how proteins are targeted to the nucleus and across membranes - Signal sequences: many protein sorting signals are localized in a stretch of aa's, can be at protein termini or on internal stretches= like a zipcode= localization seq.!- Protein transport: cytoplasm <--> nucleus!- NPC: nuclear pore complex, made of 30 nucleoporins, these pore proteins determine what prots are allowed to be imported or exported out of the nucleus!- nuclear localization signal on a protein recog. by a nuclear import receptor!- NIRs often specialized towards a subset of cargo proteins!- same idea for nuclear export!- Gated transport!- Protein translocation in mitochondria!- transmembrane transport!- mitochondria can't make everything themselves, so need to transport stuff in!- who goes where is reg. by localization seqs, protein translocator complexes!- Protein translocator complexes mediate transport across the inner and outer mitochondrial membrane!- depending on localization seq. on protein, will be integrated into the membrane or go all the way in to the matrix space !- Peroxisomes!imp. for vesicular transport- = an oxidative organelle that participates in molecule degradation to harvest energy (don't gen. ATP though, just chop things into more maneagable sizes that will then go the mitochond)!- produce H202 instead of H20 during the last electon transfer step!- Catalase: 2H202 --> 2H2O + O2!- short aa seqs target prots across the peroxisome membrane!- they get their enzymes to do stuff from cytoplasm via transmembrane transport 3) Understand the role of the ER in protein synthesis and trafficking - Protein transport into the ER - ER= netlike labyrinth!- Lipid and protein biosyn., stores intracellular Ca2+!- all secretory proteins + proteins targeted to ER lumen, golgi, or lysosomees are initially brought to ER!- going to ER, golgi, or lysosomes: translation couples to transport!- going to nucleus, mitoch, or perox: post-translational translocation!- proteins targeted for ER have ER signal seq!- SRP (signal recog. particle) binds ot ER ss and to SRP receptor!- Translocator inserts the ss into the membrane and translocates protein across the membrane!- Soluble protein transport: signal peptidase will cut out the ss, protein is freely swimming in the ER lumen!- Membrane bound protein transport: protein supposed to stay in membrane, has 2nd seq. (stop seq., usually hydrophobic aa's), this plasma membrane can bud off, but protein will stay in the membrane!- Insertion of multi-pass membrane prots --> multiple start and stop seqs (hydrophobic seqs w/hydrophilic stretches between them)!- Protein glycosylation in the ER: N-linked!- oligosacc is put onto protein while the protein is being transported into the ER lumen! 4) Understand the role of the Golgi and trans-Goli vesicular trafficking - Vesicular Transport: Transport vesicles are coated!- 3 main types of coated ves: clathrin, COPI, COPII!- ER --> Golgi COPII!- Golgi --> ER COPI!- Between golgi: COPI!- Golgi --> outside COPI!- Endocytosis: Clathrin!- Golgi --> Lysosome Clathrin!- Functions of coat: molds/shapes vesicle and [ ]s specific membrane proteins in one patch!- Phosphoionsitides mark organelles and membrane domains!- like a zipcode!- different orgs have distinct sets of PIP kinases and phosphatases!- diff. PIPs are located on diff. membrane !- has fatty acid tail in membrane connected by phosphate to carb top sticking out, can be recog. by receptors/readers!- Vesicle targeting: Rab proteins and SNARES - Protein transport from ER --> Golgi !!!!!!!!!!- Golgi: processing of oligosaccs (got them from the ER)!- diff. parts of the golgi perform distinct oligosacc processing steps!- golgi can be suubdivided (ER--> cis, medial, trans --> lysosome, pm, secretory vesicle)!- remodifying protein as it goes thru golgi!- also in golgi: O-linked glycosylation, heaviest on mucins and proteoglycans!- Lysosomes - major site for intracellular degradation of macromolecules!- contain hydrolytic enzymes the work ONLY under acidic conditions, so the lysosome is very acidic!- 3 diff. pathways deliver to lysosomes!- phagocytosis!- endocytosis!- autophagy !- specific signals get popped on in the golgi, ex. M6P signal !- lysosomal proteins get to lysosome via M6P signal and budding of vesicles ! 5) Know the basic mechs of endocytosis and exocytosis - Endocytosis - Phagocytosis: ingestion of large particles (microorganisms and dead cells) - Pinocytosis: ingestion of fluids and solutes - Receptor-mediated endocytosis: often clathrin-coated!ex. LDL particle from extracellular --> docks to LDL receptor in pm w/clathrin coated pit --> endocytosis --> uncoating --> fusion w/endosome --> turns into lysosome --> transport vesicle bud off of endosome and return LDL receptors to pm!- Exocytosis: - Constitutive secretory pathway: for proteins that are constantly produced Transport Vesicle Formation- Regulated secretory pathway - secretory vesicles are stored just underneath the pm!- they release their contents to cell exterior by exocytosis in response to a specific signal! - ex. insulin in pancreatic beta cell 6) Know the functions of the discussed organelles - See
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