TCA Cycle Flashcards
What are 5 Cofactors needed by the PDH complex? Which of these are prosthetic groups?
1) CoA-SH
2) NAD+
3) TPP (thaizole ring)
4) Lipoate arm (reduced form)
5) FAD
* *Prosthetic groups:
- Lipoate on the Lys of E2
- FAD is covalently attached to E3
What functional groups are found on Acetyl Coenzyme A? Which one serves as the “business end”?
1) Thioester (business end sulfhydryl)
2) Amine/peptide groups
3) Phosphoanhydride
4) Hydroxyl
5) Phosphoryl
6) Imidazole-like ring
7) Amino (NH2) group
How is the lipollysine arm of the PDH complex modified from lipoic acid to be used in catalysis?
The lipoate cofactor is modified from the lipoyl synthase enzyme that makes the lipoate and adds sulfur atoms that come from SAM and are synthesized by the cell
We are using the octanoic acid to covalently attach to the Lys residue E2 on the PDH complex
In what way does Acetyl CoA serve as a branch point intermediate?
1) Can enter TCA to yield energy if needed
2) Can enter FA synthesis for fat anabolism to generate storage lipids if needed
Which enzyme does lipoyllysine covalently attach to?
E2 = dihydrolipoyl transacetylase
Why is the outer mitochondrial membrane permeable to most fat-soluble small molecules and ions?
Freely permeable Porin channels
Why would you need the inner mitochondrial membrane to be impermeable to most small molecules including H+?
Need to establish a H+ ion electrochemical gradient between the intermembrane space and the inner mitochondrial matrix to create electricity driven proton pumps that will mobilize ATP sythase integral membrane protein
What 4 components would you expect to find on the inner mitochondrial membrane but NOT on the outer mitochondrial membrane?
1) Respiratory Electron Carriers (Complexes I, II, III, IV)
2) ADP-ATP translocase (brings in ATP and turns it into ADP)
3) ATP Synthase (F0F1 subunits are integral membrane proteins)
4) General membrane transporters and shuttles that are integral membrane proteins
How do we get pyruvate into the mitochondrial matrix?
Pyruvate is made in the cell cytosol by glycolysis –> outer membrane porin channel –> pyruvate/H+ symporter on the inner mito membrane
Where in the mitochondria would you expect to find Fatty acid oxidation enzymes? Amino acid oxidation enzymes? TCA cycle enzymes?
Inner mitochondrial matrix
What are the 3 enzymes involved in forming the PDH complex to convert pyruvate into Acetyl-CoA? Which one of these enzymes confers substrate specificity?
Pyruvate dehydrogenase = E1 *substrate specificity
Dihydrolipoyl transacetylase = E2
Dihydrolipoy dehydrogenase = E3
How is pyruvate (3 carbons) broken into a 2 carbon compound? Which co-factors were involved in catalysis?
CO2 is liberated from pyruvate upon
1) nucleophilic attack from TPP (deprotonated carbanion on thiazole ring) which will attack the carbonyl to 2) hydrate it to an alcohol to 3) cleave the bond and 4) generate a CO2 leaving group
In the PDH complex, how is the 2C active acetaldehyde (stabilized on TPP) then transferred to the lipoyllysine arm?
The carbanion is reformed on the 2C molecule by general base catalysis which acts as the nucleophile to attach the oxidized disulfide on the lipoyllysine arm
What was the role of TPP in the PDH complex mecahanism?
Transfer an acetyl group to the reduced lipoate arm
Catalyze decarboxylation of pyruvate by formation of a hydroxyethyl TPP
What are the sulfhydryl groups of the lipoic acid arm (reduced form) used for in the PDH mechanism to regenerate the lipoic acid disulfide bond?
Used to reduce the FAD on E3 to FADH2 with the regeneration of the lipoic acid disulfide bond
