Intracellular Compartments and Protein Sorting Flashcards
Recognize and name the membraneous organelles in eukaryotic cells
1) Nucleus
2) ER (rough and smooth)
3) Golgi
4) Endosome
5) Lysosomes
6) Peroxisomes
7) Mitochondria
8) Secretory Vessicles
Nucleus
controls cell activities
Cell Membrane
- support
- protection
- controls movement of materials in/out of cell
- barrier between cell and its environment
- maintains homeostasis
Nuclear membrane
*Controls movement of materials in/out of nucleus
Cytoplasm
*supports /protects cell organelles
ER
The ER has a central role in lipid and protein biosynthesis. Its membrane is the site of production of all the transmembrane proteins and lipids for most of the cell’s organelles, including the ER itself, the Golgi apparatus, lysosomes, endosomes, secretory vesicles, and the plasma membrane. The ER membrane makes a major contribution to mitochondrial and peroxisomal membranes by producing most of their lipids. In addition, almost all of the proteins that will be secreted to the cell exterior—plus those destined for the lumen of the ER, Golgi apparatus, or lysosomes—are initially delivered to the ER lumen.
- *Functions of the ER:
1) It is the birthplace of membrane proteins.
2) It is the site of lipid biogenesis lipids and cholesterol start in the ER and from there flow to other parts of the cell
3) It is a special environment with respect to its lumen because it is very high in calcium and thus a store house for calcium
4) It is a place where there are enzymes that are involved in lipid and hydrophobic toxin detoxification are present This is very prevalent in the liver
5) It is also this minting environment for making new proteins. Half of the proteins are manufactured into the lumen into the ER
Ribosome
*produces proteins
Mitochondria
Breaks down sugar molecules into energy, they do most of the oxidation but peroxisomes do some too!
There are about 200 mitochondria per average cell. Consider the mitochondrion to be prokaryotic endosymbiont. The inner convoluted membrane can be considered as the membrane of the endosymbiont. The outer membrane could be considered as the vestiges of the membrane that originally engulfed the symbiont. Thus there are four compartments, matrix, inner membrane, inter-membrane and outer membrane.
Mitochondria have their own DNA genome and make some of their own proteins. But two billion years of symbiotic relationships have created interdependencies, such that many mitochondrial proteins are encoded by the nuclear genome. Hence the evolution of systems for getting proteins from cytosol into mitochondria.
Vacuole
*store food, water, waste (plants need to store large amounts of food)
Lysosome
- breaks down larger food molecules into smaller molecules
* digests old cell parts
Peroxisomes
Peroxisomes originally were defined as organelles that carry out oxidation reactions leading to the production of hydrogen peroxide. Because hydrogen peroxide is harmful to the cell, peroxisomes also contain the enzyme catalase, which decomposes hydrogen peroxide either by converting it to water or by using it to oxidize another organic compound. A variety of substrates are broken down by such oxidative reactions in peroxisomes, including uric acid, amino acids, and fatty acids. The oxidation of fatty acids (Figure 10.25) is a particularly important example, since it provides a major source of metabolic energy. In animal cells, fatty acids are oxidized in both peroxisomes and mitochondria, but in yeasts and plants fatty acid oxidation is restricted to peroxisomes.
Functions of Peroxisomes:
1) Oxidation of very long chain fatty acids
2) Oxidation of branched chain fatty acids
3) Oxidation of cholesterol to bile acids (liver)
3) Synthesis of plasmalogens (membrane lipids found in myelin)
4) Oxidation (detoxification) of some metabolic intermediates and foreign substances for elimination (flavoprotein oxidases and catalase)
5) Decomposition of hydrogen peroxide
Compare a signal peptide to a signal patch
Signal peptides are one sequence where the signal patches are distant to one another but come close to each other once the protein has folded.
Recognize features of signal peptides that direct proteins to specific compartments
1) ER signals have a hydrophobic center.
2) Mitochondrial signals are amphipathic alpha helices
3) Nuclear import signals are very basic
4) Perosixomal signals are SKL-COOH (serine, lysine, leucine)
5) Nuclear export signals are leucine-rich
ER signals
ER signals have a hydrophobic center.
Mitochondrial signals
Mitochondrial signals are N-terminal amphipathic alpha helices
Nuclear import signals
Nuclear import signals are very basic
Perosixomal signals
Perosixomal signals are SKL-COOH (serine, lysine, leucine)
Nuclear export signals
Nuclear export signals are leucine-rich
Identify specific organelles that import proteins from the cytosol after synthesis of the protein is completed (post-translationally)
1) Nucleus- Uses Ran-GAP (dephosphorylates to get Ran-GDP in cytosol), Ran-GEF (switches of GDP for GTP to get Ran-GTP in nucleus), Ran-GDP, and Ran-GTP. There is also a nuclear import or export receptor that binds to Ran-GTP or GDP.
2) Mitochondria- Uses the TOM and TIM23 complexes to feed unfolded protein into mitochondria. Also uses the help of cytosolic Hsp70 to keep it unfolded and mitochondrial Hsp70 and Hsp60 to refold protein. The N-terminus amphipathic alpha helix is the
3) Peroxisomes
ER IS CO-TRANSLATIONALLY
Identify nuclear localization signals and their importance in protein import in the nucleus.
Nuclear export signals are leucine-rich. It MUST contain a stretch of Leucines (Leu or L). If one is mutated, it will remain in the cytosol. These are important to bring the necessary proteins into the nucleus. Without these signal, the proteins would have no way of entering.
Compare and contrast protein transport into (and out of) the nucleus with transport into other organelles, with regard to conformation of protein, the dimensions of the pore, the requirement for energy, and the subsequent fate of the import or export signal
1) Nucleus- Uses Ran-GAP (dephosphorylates to get Ran-GDP in cytosol), Ran-GEF (switches of GDP for GTP to get Ran-GTP in nucleus), Ran-GDP, and Ran-GTP. There is also a nuclear import or export receptor that binds to Ran-GTP or GDP. Here the fully folded protein is transported into the nucleus and thus the nuclear pore is much larger. The nuclear localization signals STAY ON because the nucleus breaks down and reforms after cell division or mitosis. They need to know where to go again. This way the cell is not having to remake protein! This requires GTP to be moved.
2) Mitochondria- Uses the TOM and TIM23 complexes to feed unfolded protein into mitochondria. Also uses the help of cytosolic Hsp70 to keep it unfolded and mitochondrial Hsp70 and Hsp60 to refold protein. The N-terminus amphipathic alpha helix is the signal. It is fed into the mitochondria BEFORE folding using Hsp70 to assist this. There is a requirement for ATP TWICE, once at TOM and once at TIM23 because the Hsp70 are ATPases that require ATP to ratchet it in. The signal peptide is cleaved because the protein will remain there! Smaller pore.
3) Peroxisomes- They acquire some proteins via budding so they are fully folded proteins. Other proteins are also introduced post-translationally when folded. Larger pore.
4) ER- Proteins are inserted CO-TRANSATIONALLY using a SRP (signal recognition particle) that binds to a receptor on the ER. The pore is thus relatively smaller to fit a chain into it. This process using the Sec61 translocon. It is not using ATP, rather the ribosome is pushing it in.
Describe the roles of Ran, the accessory proteins Ran-GEF and Ran-GAP, and import and export receptors in regulated transport through the nuclear pore
Ran-GDP is in excess in the cytosol and Ran-GTP is in excess in the nucleus. When a protein binds to a nuclear import receptor, it will be taken into the nucleus. In order to dissociate from the nuclear import receptor a Ran-GTP needs to bind. This causes the protein to be dissociated and then the Ran-GTP bound nuclear import protein will be exported. Once it gets into the cytosol, Ran-GTP will dissociate from the nuclear import protein. The Ran-GTP will then quickly encounter at Ran-GAP in the cytosol will then convert the Ran-GTP to Ran-GDP.
When a protein needs to to be exported from the nucleus, it will encounter a nuclear export receptor that binds to its signal. It will also bind Ran-GTP. It will then be exported from the nucleus and then when the Ran-GTP is dephosphoylated via a Ran-GAP, the protein dissociates.\
Ran-GEF is essential in reestablishing Ran-GTP from Ran-GDP that is produced in these two steps but exchanging a GDP for GTP
Explain how the regulation of the transport of transcription factors through the nuclear pore can lead to changes in the expression of specific genes
If the transcription factors are being shuttled in with a nuclear import signal, then there will be an increased in transcription. However, if the transcription factors have a nuclear export signal that develops, they will be signaled for export and gene expression will thus be decreased.
Identify the components of the mitochondrial import machinery and their functions. Compare the functions of chaperone proteins (heat shock protein families, Hsp60 and Hsp70), ATP and electrochemical energy in mitochondrial protein import.
Uses the TOM and TIM23 complexes to feed unfolded protein into mitochondria. Also uses the help of cytosolic Hsp70 to keep it unfolded and mitochondrial Hsp70 and Hsp60 to refold protein. The N-terminus amphipathic alpha helix is the signal. It is fed into the mitochondria BEFORE folding using Hsp70 to assist this. There is a requirement for ATP TWICE, once at TOM and once at TIM23 because the Hsp70 are ATPases that require ATP to ratchet it in. The signal peptide is cleaved because the protein will remain there! Smaller pore.
So, an UNFOLDED peptide with a signal sequence associates with TOM and Hsp70 chaperones. ATP is required to ratchet it into the intermembrane space. Then it goes to TIM23 and mitochondrial Hsp70 on the INSIDE of the mitochondria help pull it through once it gets going via electrophoretic processes (inside is negative and the N-terminus of peptide is + so it wants to go in). It requires ATP because the Hsp70s are ATPases. There are also Hsp60s that are in the mitochondria and take in the protein in a barrel like structure to help it fold in the mitochondria like Hsp70.
