Intracellular Compartments Flashcards
What are nuclear localization signals (NLS)?
Short stretches of amino acid sequences on proteins
(1) What are importins? (2) What role do they play in protein transport? (3) Describe its structure.
(1) Specialized nuclear import receptors
(2)
• Recognize NLS of cargo proteins and transport them through nuclear pore complexes (NPCs)
(3) Structure:
• Alpha-subunit - binds NLS of cargo protein to form cargo-receptor complex
• Beta-subunit - binds cytoplasmic fibrils and docks cargo-receptor complex onto channel’s opening
Explain how the Cargo-receptor complex passes through a nuclear pore.
• Beta subunit makes multiple weak contacts with FG repeats and hops across channel
What happens inside the nucleus after the protein has been imported?
• Ran-GTP - binds to importin beta —> induces conformational change —> importin releases cargo protein
• Importin-Ran-GTP complex is transported back to cytosol
• GTP is hydrolyzed
What is the role of SRP in protein transport?
• SRP - binds to signal sequence at N terminal
• Structure:
—> Signal sequence binding pocket
—> Translation pause domain - blocks elongation factor binding site on ribosome and arrests translation
—> GTP-binding domain
• Binds to ribosome-nascent chain complex —> changes conformation of SRP (exposes receptor binding site)
• Docks at SRP receptor on ER membrane
• SRP-SRP receptor complex carries ribosome and polypeptide chain to translocon channel
• RNC complex is unloaded on translocon
• GTP hydrolysis dismantles SRP-SRP receptor complex to recycle
(1) What is cotranslational translocation? (2) What happens during this process?
(1) Process where proteins enter ER lumen while still under synthesis
(2)
1. SRP-SRP receptor complex unloads ribosome-nascent chain complex on Sec61 channel
- Signal sequence at N terminal latches onto signal sequence Recognition site
- Signal sequence exists through lateral gate of channel to enter signal peptidase complex (cleaves off signal sequence)
- ER chaperones (e.g., BiP) bind translocated protein and assist in protein folding
Explain the insertion of single-pass transmembrane proteins in RER. In other words, how is the process of cottanslational translocation different in transmembrane proteins?
- ER signal sequence of transmembrane protein - acts as start-transfer signal
- Signal sequence is cleaved off by signal peptidase complex —> release N terminal into lumen
- Hydrophobic domain in polypeptide chain acts as stop-transfer signal (cannot cross Er membrane) —> forms transmembrane domain
- Ribosome continues synthesis of cytosolic domain
What is the difference between Type I and Type II proteins?
Type I protein:
• N terminal in lumen
• C terminal in cytosol
Type II protein:
• Hydrophobic domain preceded by positively charged residues —> N terminal remains in cytosol
• C terminal in lumen
• “Upside-down type I protein”
What are the three categories of membrane trafficking? Explain each of them.
- Secretory pathway - cargo is transported within cell from organelle to another
• Substances are packaged into vesicles that can be transported from one organelle to another - Endocytosis - cargo is transported into cell
- Exocytosis - cargo is transported out of cell
• Vesicles fuse with plasma membrane, releasing cargo into extracellular space
What are some substances that need to be exported via exocytosis?
• Waste products
• Membrane proteins
• Signaling molecules
What are some substances that are not produced in the cell and are therefore imported via endocytosis?
• Vitamins
• Cholesterol
• Micronutrients
Explain two types of endocytosis.
- Pinocytosis -
- Phagocytosis -
(1) What are coated vesicles? (2) What are the three types?
(1) Coated vesicles - transport vesicles that bud off from specialized regions of the cell membrane; specific coat proteins cover their surface
(2)
1. COPI vesicle
• Transport molecules between different parts of Golgi body and from Golgi back to RER
• Composed of coatomers
- COPII vesicle
• Formed in ER membrane
• Transport from ER to Golgi
• Composed of coatomers - Clathrin-coated vesicle
• Transport proteins from Golgi to plasma membrane
How are clathrin-coated vesicles formed?
- Clathrin - protein that forms outer layer of coat
- Adaptor protein - forms inner layer of coat
- Cargo and receptor are packed into a clathrin-coated bud
- Dynamin (GTP-binding protein) - attaches around neck of bud —> triggers GTP hydrolysis —> drives conformational change in dynamin —> neck of bud stretches until vesicle pinches off
For a vesicle to fuse with the target organelle, the lipid bilayers of the two membranes must be within _______ of each other
1.5 nanometers
What are SNAREs? How do they fuse membranes?
• Transmembrane proteins with helical motifs that catalyze membrane fusion
• v-SNARE
• t-SNARE
• Helical domains of v-SNARE and t-SNARE wrap around one another —> forms trans-SNARE complex (initiates membrane fusion)
•
What is the function of the NSF?
• hexameric ATPase
• Catalyzes SNARE disassembly
• Dissociates v- and t-SNAREs
Protein destined for ER have a _______ signal sequence at their N terminal.
Hydrophobic
