sakai-PDH and TCA cycle Flashcards
How is pyruvate transported from the cytosol into mitochondria? Where in the cell are the PDH complex and the TCA cycle performed?
Pyruvate in the cytosol is transported by a pyruvate carrier across the inner mitochondrial membrane. In this process pyruvate enters together with H+ in exchange for OH- .
The PDH complex and the TCA cycle are both performed inside of mitochondria.
The pyruvate dehydrogenase complex (PDH) in mitochondria allows the connection of aerobic glycolysis with the TCA cycle. What is the overall reaction?
Pyruvate and free CoA (HSCoA) and NAD+ are used to form acetyl CoA and NADH and carbon dioxide.
Describe the actions of enzyme 1, 2 and 3 of the PDH complex.
Enzyme 1 decarboxylates pyruvate to a molecule that is used as substrate for enzyme 2.
Enzyme 2 uses HSCoA and forms acetyl CoA and carbon dioxide.
Enzyme 3 uses NAD+ and forms NADH.
What are the coenzymes (cofactors) of the individual enzymes of the PDH complex? Are they also needed in other pathways? Summarize!
E1 needs thiamine-PP (from vitamin B1) .
E2 needs lipoic acid. It uses HSCoA and forms acetyl CoA.
E3 needs FAD and uses NAD+ and forms NADH.
What coenzyme is needed for TCA (alpha keto glutarate DH), PPP(transketolase), and PDH(enzyme 1)?
Thiamine-PP
when Thiamine-PP is absent what difiecency occurs?
Deficiency in Wernicke-Korsakoff and in Beri-Beri
Thiamine-PP is formed from vitamin B1.
what coenzyme is needed for PDH: enzyme 2,
TCA: -ketoglutarate DH?
Lipoic acid
what is special about lipolic acid?
Inhibition by trivalent arsenic (arsenite)
what coenzyme is needed for PDH: enzyme 2
TCA: -ketoglutarate DH,-oxidation of fatty acids?
HSCoA
what is special about HSCoA?
Acetyl CoA and succinyl CoA contain energy rich bonds
what coenzyme is needed for PDH: enzyme 3
TCA: -ketoglutarate DH, and succinate DH?
FAD
what is sepcial about FAD?
FAD is a prosthetic group and is tightly bound to the enzyme.
FADH2 leads in the ETC and oxidative phosphorylation to 2 (1.5) ATP
what coenzyme is needed for Glycolysis: glyceraldehyde 3-P DH PDH: enzyme 3 TCA: isocitrate DH -ketoglutarate DH malate DH?
NAD+ is a free coenzyme found in cytosol and mitochondria
NADH leads in the ETC and oxidative phosphorylation to 3
(2.5) ATP
PDH is an important link between aerobic glycolysis and the TCA. How is it regulated in general terms?
PDH is product inhibited by acetyl CoA and NADH.
Calcium ions stimulate PDH. They activate PDH phosphatases.
NADH is formed in E3, and acetyl CoA is formed in E2.
[Acetyl CoA and NADH formed by -oxidation and the NADH generated in the TCA cycle also lead to inhibition of PDH.]
PDH is also inhibited by phosphorylation by a specific PDH kinase. How is this PDH kinase allosterically regulated?
PDH kinase phosphorylates and inhibits PDH at enzyme 1 (pyruvate decarboxylase).
PDH kinase is allosterically activated by acetyl CoA and NADH and ATP.
PDH kinase is allosterically inhibited by pyruvate.
That means that when ATP is high, and acetyl CoA and NADH are high, the PDH should not be used to form acetyl CoA and pyruvate shall be saved.
When do we want to activate PDH? Describe the action of calcium in regulation of pathways! Discuss the TCA cycle and glycogen degradation!
The PDH shall be active during muscle contraction when glycogen degradation and aerobic glycolysis lead to pyruvate that can be used as acetyl CoA in the TCA cycle.
In general, calcium ions activate
glycogen phosphorylase kinase using the calmodulin subunits,
the PDH complex (indirect via activation of PDH phosphatase)
and the TCA at the level of isocitrate DH and -ketoglutarate DH.
Are PDH kinase and PDH phosphatase mainly regulated by hormones or are they mainly allosterically regulated?
They are mainly allosterically regulated by the energetic demands inside of the cell.
Calcium ions activate PDH phosphatases and leads to an active PDH complex.
(Insulin may act via enzyme induction of PDH phosphatases)
Describe PDH regulation in the liver!
In the liver, PDH has to be tightly regulated, as after pyruvate is used to form acetyl CoA, the carbons of the acetyl-group cannot be used for gluconeogenesis.
In the liver, pyruvate is saved for gluconeogenesis when needed.
The PDH complex is inhibited by acetyl CoA and NADH generated by -oxidation during fasting.
[Pyruvate carboxylase is used for gluconeogenesis and forms oxaloacetate. It needs the absolute activation by acetyl CoA which is formed during -oxidation of fatty acids]
Which enzyme of the TCA cycle needs the same coenzymes as the PDH complex? Name other similarities!
The TCA cycle enzyme -ketoglutarate DH needs the same coenzymes as the PDH complex.
Both are inhibited by NADH and both are activated by calcium ions, but in a different way.
Both catalyze a decarboxylation, use HSCoA, and form NADH.
PDH forms acetyl CoA, TCA cycle forms succinyl CoA.
Special for PDH and not found for ketoglutarate DH is that PDH is also regulated by PDH kinase and PDH phosphatase.
PDH kinase inhibits PDH and PDH kinase is activated by acetyl CoA and NADH.
PDH phosphatase activates PDH and is itself activated by calcium ions
Describe congenital lactic acidosis. Which enzyme of the PDH complex is deficient?
Enzyme 1 is deficient and the pyuvate is used to form lactate.
Symptoms include neurodegeneration, muscle spasticity and can lead to early death.
In case of poisoning with trivalent arsenic (arsenite), which coenzyme is bound by arsenite and inhibited? Which enzyme of the PDH complex is inhibited? Which enzyme of the TCA cycle is inhibited?
Arsenite forms a stable complex with lipoic acid.
Lipoic acid it used in E2 of the PDH complex, and also by -ketoglutarate DH
Compare poisoning with trivalent arsenic to the poisoning with pentavalent arsenic (arsenate). Which enzyme of glycolysis uses arsenate instead of inorganic phosphate?
Trivalent arsenic (arsenite) binds to lipoic acid. Pentavalent arsenic (arsenate) competes with inorganic phosphate.
Arsenate is used instead of inorganic phosphate by glyceroaldehyde 3-phosphate dehydrogenase.
Instead of the formation of 1,3-bisphosphoglycerate, a now instable molecule is formed which immediately leads to 3-phosphoglycerate instead of 1,3-bisphosphoglycerate.
With that, one of the two steps of substrate level phosphorylation in glycolysis cannot be performed normally
Discuss the consequences of thiamine deficiency (Wernicke-Korsakoff and Beri-Beri) related to PDH, TCA cycle and PPP.
Brain is dependent on energy generation by aerobic glycolysis, PDH and TCA cycle.
Thiamine-PP is needed for the PDH and -ketoglutarate DH. This means that deficiency of thiamine-PP leads to less generation of ATP.
In cells with high levels of NADPH, the oxidative part of the PPP cannot be used to form ribose-5-P. Transketolase fulfills the important function to generate ribose-5-P (together with transaldolase in the reversible part of the PPP) .
Transketolase is two times involved and needs thiamine-PP as coenzyme. The deficiency of thiamine-PP leads to less formation of ribose 5-P in cells that cannot use the oxidative part of the PPP.
Explain the central role of the TCA cycle related to catabolism and anabolism.
The TCA cycle has catabolic functions including two steps of decarboxylations.
It is also able to generate 3 NADH and 1 FADH2 and 1 GTP.
On the other hand, the TCA cycle has also anabolic function, for example related to synthesis of amino acids or of heme (succinyl CoA) . It also provides metabolites from glucogenic amino acids which can be used for gluconeogensis in the liver.