Protein Metabolism - Regulation Flashcards
What is the goal of the Lac Operon/When do you want it to work?
The lac operon wants to metabolize lactose (degrade it) if it is the only preferred fuel source
What does the presence of lactose do to the repressor [Lac Operon regulation]
B-gal converts lactose into allolactose. Allolactose inhibits the repressor from binding. Thus, the genes are translated.
What does the presence of glucose do to the Lac Operon [Lac Operon Regulation]
Summary: no glucose, yes camp, yes crp, yes translation (vice versa)
Explanation: glucose steals the phosphate groups from a series of enzymes that need to be phosphorylated in
What is the goal of the tryptophan operon?
The tryptophan operon wants to make tryptophan when the concentrations of tryptophan drop
How does the repressor function depending on tryptophan concentration [TRP operon regulation mechanism]
In the presence of tryptophan, repressor binds, no translation
No tryptophan, no binding of repressor, translation
Transcriptional Attenuation [TRP operon regulation mechanism; What happens at low and high tryptophan concentration?]
Transcriptional Attenuation depends on availability of tryptophan charged tRNA and position of the ribosome
Position of ribosome is controlled by two adjacent TRP codons. TRP codons place 2 adjacent tryptophans on a leader peptide.
At high [TRP], ribosome synthesizes leader peptide with no problem. Continues over regions 1 and 2, so 3-4 stem loop forms which is before poly U sequence. Terminates.
At low [TRP], ribosome cannot synthesize leader peptide easily. Ribosome stuck only on region 1, so 2-3 stem loop forms, which does not have a poly U sequence. No termination, Translation continues.
4 Iron Homeostasis Proteins
1) Transferrin (import)
2) Ferritin (storage)
3) Aconitase from citric acid cycle (ex. Of usage)
4) Fpn1 (export)
Iron Response Element @ 3’-UTR/Which Protein is regulated this way [Consider which protein you want translated depending on the environment]
-Presence of iron, IRP falls off and exposes termination sequence, and gene stops being translated
-no iron, no exposure of termination sequence, gene is translated
Protein: Transferrin. When there’s no iron, you want more iron to be imported
Iron Response Element @ 5’-UTR/Which Protein is regulated this way [Consider which protein you want translated depending on the environment]
-Presence of iron, IRP falls off and gene is accessible
-No iron, irp bound and gene is inaccessible
Protein: Ferritin. When there is iron, you can store this iron without starving the cell.
R-Protein Operon [Goal of operon? ]
The goal is to stop ribosomal protein synthesis if there is not enough rRNA
R-Protein Operon [Which has higher Kd mRNA or rRNA? Which has higher affinity?]
Kd, mRNA > Kd, rRNA. Thus, r-proteins have greater affinity for rRNA.
R-Protein Operon [Which Protein is involved?]
Protein L4
Translation Riboswitch [How does it work/ When does it work?]
When ligand is present, a secondary structure forms and includes the shine-delgarno sequence, which sequesters it away from the ribosome
What happens when elf2 is phosphorylated? What is the name of the kinase and what regulates the kinase [globin protomer synthesis regulation]
When elf2alpha is phosphorylated by kinase HCR, it forms a stable complex of elf2alpha and elf2B. This complex does not translate and does not make globin
heme inhibits kinase and translation is active. Makes sense because if there is heme there needs to be protein.
How proteins get into the ER (STEPS DO NOT CORRESPOND TO IMAGE)
1) mRNA is being read by ribosome
2) Signal sequence recognition protein (SRP) binds to the signal sequence . Translation stops because of the bound SRP. (SRP binds stops translation)
3) SRP protein binds to membrane protein on the outer surface of the ER (SRP receptor). This opens a channel called the peptide translocation complex and the peptide chain from the ribosome gets threaded into the ER. (SRP binds receptor and threads protein through channel)
4) SRP protein dissociates because of a GTP timer, and translation can continue (SRP leaves translation continues)
5) Protein is further threaded into the ER
6) Signal sequence is cleaved by a signal peptidase (signal cleaved)
7) Ribosome dissociates from the ER
How does Glycosylation happen? [Soluble proteins]
-To glycosylate a protein, carbohydrates are attached to a molecule of dolichol phosphate
-Phosphate of dolichol phosphate points to the cytosol and dolichol points into the ER
1) UDP versions of carbohydrates are activated sugars. Carbohydrates are added to the dolichol phosphate. This step is inhibited by tunicamycin
2) More carbohydrates are attached (adding)
3) Translocation occurs. The dolichol is now facing the cytosol and the phosphates attached to the carbohydrates are facing into the ER. (flipping)
4) More carbohydrates are added
5) Asparagine side chain nitrogen is the nucleophile and the entire glycan is transferred. Dolichol phosphate leaves (protein takes glycan)
6) Molecule of dolichol pyrophosphate is translocated and a phosphate group leaves thus recycling it into dolichol phosphate (dolichol pyrophosphate recycled)
How are membrane proteins inserted? What gets stuck in the translocon? [N-terminus inside]
- N-terminus points into the ER lumen and the peptide is threaded through a channel called a translocon
- Movement through the translocon stops when an internal stop sequence (hydrophobic helix) gets stuck within the translocon. This happens because the translocon can only move polar segments all the way through into the lumen.
How are membrane proteins inserted? What gets stuck in the translocon? [C-terminus inside]
- This polypeptide chain features an internal signal sequence that cannot be cleaved by a peptidase and remains in the translocon
- Forms a hairpin like shape because the internal signal sequence gets stuck in the translocon
- Ribosome extends the peptide on the C-terminal side and threads it through
How do proteins get into the nucleus? [3 steps]
1) two cytosolic proteins (importin-alpha and importin beta) bind to the nuclear localization sequence
2) Complex of the three proteins binds to the nuclear pore complex which catalyzes hte ATP dependent movement into nucleus
3) Importans fall off and protein gets dropped off in the nucleus
How are the importins recycled? (how are importins brought back from nucleus to cytosol) 3 steps
1) RAN GTPase binds to the beta importin
2) CAS (cellular apoptosis susceptibility) and RAN GTPase proteins binds to alpha importin
3) Both alpha and beta complex leave the nucleus and with GTP hydrolysis they are dropped off in the cytosol
How is the RAN-GDP made functional again?
Meanwhile, Ran-GDP now re-enters the nucleus by binding to nuclear transport factor-2 (NTF2). Once in the nucleus, a specific GEF protein catalyzes the exchange of GTP for GDP, causing NTF2 to be removed from Ran-GTP.
How are proteins ubiquinated?
1) ubiqutin gets an adenylyl group added to it (addition of leaving group)
2) E1 displaces leaving group
3) E2 displaces E1
4) E3 displaces E2
5) C-terminus becomes attached to the target protein via a side chain amino group on a lysine present on the target protein
High Glucose, Low Lactose [What happens to Lac Operon?]
lac operon off because lac repressor is bound to operator and no C(RA)P protein is bound to to the DNA
Low Glucose, Low Lactose [What happens to Lac Operon?]
lac operon off because lac repressor is bound. (if there is no lactose, there is no allolactose)
