Endoplasmic Reticulum Flashcards
What is endoplasmic reticulum, how is it divided and what are their functions?
Continuous net of branching tubules and flattened sacs responsible for the synthesis of proteins and lipids. Rough ER contains ribosomes and synthesizes proteins and smooth ER is a transition cisternae and specialized site for vesicular transport to Golgi aparatus. rER is continuous with the nuclear envelope which is sudded with ribosomes on the outer side as well.
Transmembrane or water soluble protein synthesis for:
- ER itself
- Golgi
- lysosomes
- endosomes
- secretory vesicles
- peroxisomes
- plasma membrane and other membranes (like mitochondria)
What is the function of sER?
sER is a transitional compartment and an exit site for vesicular transport to Golgi. It is highly abundant in some cell types.
Specialized functions:
- lipid metabolism (hepatocytes)
- steroid hormone synthesis
- detoxification using P450 (hepatocytes)
- Ca2+ storage (sacroplasm, muscle cells)
Where are the ribosomes synthesized and what is their structure in prokaryots and eukaryots?
The ribosomal subunits are synthesized on the nucleous organizing centers on certain chromosomes, exit the nucleus via nuclear pores and only then are assembled to prevent undesired translation.
Structure
P:
50S + 30S (70S together) with respective proteins and rRNA within them (L: 31 proteins and S:21 proteins; 23S and 5S in the large subunit and 16S in the small subunit)
E:
60S + 40S (80S together) with respective proteins and rRNA within them (L: 50 proteins and S:33 proteins; 28S, 5.8S and 5S in the large subunit and 18S in the small subunit)
Define the difference between free ribosomes and rER bound ribosomes.
Structurally identical. Ribosomal subunits are sourced from a free pool of ribosomes that is available for the cells’ use. Based on the type of mRNA, they either stay in the cytosol and translate proteins where they create a long polyribosomal chain and slide on the mRNA or are recruited onto the membrane of the rER where the mRNA concentrates and the ribosomes again slide on the mRNA whereas they are also attached to the rER membrane to facilitate rER insertion. The translation occurs in 5’->3’ direction.
What types of rER translocation there are and with with ORG is it associated? Briefly define them.
1) Co-translational translocation in bacteria, archea, eukaryotes
Signal recognition particle (SRP) and SRP receptor form a tight seal with Sec61 translocator to prevent leakage/entering pf undesired molecules and push the protein in. No additional energy uptake needed.
2) Post-translational translocation in eukaryotes
Sec61+Sec62+Sec71+Sec72 work together with BiP molecules which are attaching on the polypeptide chain and push the protein in. ATP->ADP
3) Post-translational translocation in in bacteria
SecATPase
Do other organells have co-translational translocation except the rER?
No. Only the rER has bound ribosomes.
Explain the co-translational translocation of proteins into the rER. What two proteins play an important role in it?
mRNA that carries a specific fragment encoding for a hydrophobic signaling sequence localises close to the rER mebrane and recruits ribosomes. The ribosomes slide onto the mRNA sequence and synthesize the polypeptide chain. The specific hydrophobic sequence gets recoginsed by a signal recognition particle (SRP).
SRP complex consists of 6 subunits and 1 srpRNA and has a site for binding the hydrophobic signaling sequence of the growing protein chain and site for binding a respective receptor. Upon binding to the SRP receptor on the rER, translation is paused. Then, the ribosome is docked onto the SRP receptor and alligned with a Sec61 translocator which opens and forms a tight seal. The partially-synthesized protein enters rER via the opened translocator where a signaling peptidase cleaves off the signaling sequence of the protein. Translation is resumed and the insertion of the protein continues. SRP has a specialized binding pocket lined with methionin (unbranched and flexible therefore can bind to many sequences and shapes)
Sec61 is a plug shut translocation which aligns with ribosomes to facilitate the entry of a desired protein to the rER. It has an aquaeous pore and consist from 3/4 complexes with 3 transmembrane protein per each. No leakage occurs.
Upon insertion, the completed protein within the rER matures and ribosomes de-assemble.
How are proteins integrated into the membrane of ER? What is an example of a membrane-bound ER protein?
Proteins contain specific start and stop sequences whic indicate the insertion pattern and passing of the protein through a membrane. Proteins can have simple insertion patterns like 2-transmembrane proteins or serpentine proteins which pass the membrane 7 times.
On N-terminus of the protein, the ER insertion signal usually acts as a start sequence where a part of the peptide is inserted into the lumenal environment followed by an insertion of a stop sequence on the C-terminus. Both the N- and the C-terminus are on the cytosolic side.
For a more complex proteins like rhodopsin, the N-terminus has a stop sequence which forces its insertion into the lumenal side. The sequences then alternate between start and stop signals and result in the C-terminus to be exposed to the cytosolic side.
Protein-disulfide isomerase oxidises the SH groups to form disulfide bridges in the cysteines. This process does not normally occur due to the reducing cytosolic environment.
How are proteins glycosilated in the rER?
Proteins in the rER undergo partial glycosilation where a precursor with many sugars is synthesized. These proteins have a net of glucose, mannose and N-acetylglucoseamine attached to a specific asparagine AA which is located in a sequence of either Asn-X-Ser or Asn-X-Thr where X is any AA except proline. The sugars are attached via N-link to the protein. Afterwards, the excessive sugars are trimmed in the Golgli and the protein is left with a 5-sugar core consisting of N-acetylglucoseamine and mannose.
The sugar chain is synthesized on a carier molecule termed dolichol which is embedded in the membrane of the rER.
1) Using an energy input, a pyrophosphate macroergic bridge is gradually attached togetger with sugars in an orderly fashion on the cytosolic side. The bridge primes the potential transfer of sugars from dolichol to the asparagine
2) The dolichol molecule is flipped using transporter protein onto the lumenal side and more sugars are added from donor molecules
Glucose donor dolichol-P-glucose (membrane embedded)
-dolichol phosphate
-UDP-glucose
Mannose donor dolichol-P-mannose (membrane embedded)
GDP-mannose
3) The sugar chain is transferred to the asparagine of respective protein
4) 3 glucoses and 1 mannose is removed (still in the rER)
How are cytosolic proteins glycosilated?
Simpler glycosilation. Single N-acetylglucoseamine group added to serine or threonine.
How are proteins in the rER checked for a proper fold and folded?
1) Chaperone calnexin binds to a one terminal glucose of an incompletely folded protein, traps it in the ER.
2) Glucosidase releases the protein by cleaving the excesive terminal glucose.
3) Glucosyl transferase determines the fold and if it is badly folded it adds again a glucose molecule from UDP-glucose and the protein is reshaped once again the process repeats untill correct.
What happens to a missfolded protein from rER, unable to be correclty folded?
Same process for both soluble and transmembrane proteins.
1) Chaperones guide the protein via their respective translocator out to the cytosol.
2) Deglycosilation with N-glycanase.
3) Ubiquitylation and proteasome degradation.
How is phosphatidylcholine synthesized? For which other phospholipids is the process similar? How does the plasma membrane and the rER mebrane differ when it comes to composition? How are the phospholipids delivered to the plasma membrane
fatty acyl CoA + glycerol-3-phosphate + CDP-choline
Fatty acyl CoA gets embedded into the membrane onto the cytosolic side. The mature phospholipids are redistributed to specific membranes. Phosphatidylserine, phosphatidylinositol and phosphatidylethanolamine are synthesized in a similar fashion.
Phospholipid are redistributed differently in the plasma membrane and in the ER membrane. For ER, phopspholipids are evenly redistributed and mixed across the membrane. For plasma membrane, phospholipid are rather asymetrical. Scramblase and flippase work respectivelly based on the specific membrane type.
Transport of phospholipid takes place down the concentration gradient so no E is required (ER = higher c than plasma membrane because they are heavily synthesized there). They however need chaperone proteins because they are not soluble.
