Gluconeogenesis Flashcards
What are the roles of gluconeogenesis?
To produce glucose when blood glucose levels are low (i.e., fasting, glycogen store depletion)
Is gluconeogenesis an anabolic or catabolic pathway?
Anabolic, meaning that it requires energy, is reductive, and is divergent
What are the carbon sources for gluconeogenesis?
Pyruvate, lactate, glycerol, and some amino acids (glucogenic)
Why is gluconeogensis simply NOT the reverse reactions of glycolysis?
- Glycolysis itself is irreversible because some steps are irreversible
- Need for method of regulation
Which reactions are different in gluconeogenesis (as compared to glycolysis)?
Steps 1, 3, and 10 of glycolysis
Where does gluconeogenesis primarily occur in mammals?
In the cytosol of hepatocytes in the liver
What is the first bypass reaction in gluconeogenesis?
Pyruvate is transported into the mitochondrial matrix via pyruvate transporter and acted upon by pyruvate carboxylase with the help of biotin, its covalently attached coenzyme
Pyruvate is produced in the cytosol, but the first bypass reaction of gluconeogensis occurs in the mitochondrial matrix. How does this occur?
Pyruvate moves through porins on the outer mitochondrial membrane into the mitochondrial intermembrane space and is transported across the inner mitochondrial memberane into the mitochondrial matrix via the pyruvate transporter
Once in the mitochondrial matrix, what happens to pyruvate?
It is acted upon by pyruvate carboxylase and biotin, where it is converted into oxaloacetate
The product of catalysis by pyruvate carboxylase is oxaloactetate, which must be moved back into the cytosol to continue gluconeogensis. How does the cell accomplish this?
Through the malate-oxaloacetate shuttle system
How does the malate-oxaloacetate shuttle system function?
- Oxaloacetate is converted to malate via malate dehydrogenase
- Malate is transported out of the mitochondria via the malate transporter
- Once in the cytosol, malate is converted back into oxaloacetate by malate dehydrogenase
Malata dehydrogenase is part of what class of enzymes?
Oxidoreductase
To convert oxaloacetate into malate, what coenzyme is required?
NADH
To convert malate into oxaloacetate, what coenzyme is required?
NAD+
How many active sites does biotin have?
Two - in one active site, biotin picks up carbon dioxide; in the other active site, biotin transfers carbon dioxide
How is biotin covalently linked to pyruvate carboxylase?
Via an amide linkage at a lysine residue
What reactant is required for pyruvate to be converted to oxaloacetate via pyruvate carboxylase?
Carbonate
Is the conversion of pyruvate to oxaloacetate via pyruvate carboxylase energy-requiring or energy-producing?
Energy-requiring; it requires ATP
What is the mechanism for pyruvate carboxylase?
- The oxygen atom on bicarbonate acts a nucleophile, attacking the electrophilic phosphorus of the gamma phosphate on ATP, releasing ADP and a proton (from bicarbonate)
- The resulting molecule is an “activated molecule of carbon dioxide”
- The nitrogen atom in biotin now functions as a nucleophile and attacks the carbon atom of the carbonyl group, ultimately “kicking out” phosphate, which is a good leaving group
- A hydrogen atom from pyruvate’s methyl group deprotonates, forming a carbanion that attacks the carbonyl carbon of carboxy-biotin, ultimately kicking out biotin and reforming the enzyme by protonating nitrogen on its way out
Once oxaloacetate moves back into the cytosol it is acted up by which enzyme?
PEP carboxykinase
Oxaloacetate is converted to _________________________ via PEP carboxykinase.
Phosphoenolpyruvate
What is the mechanism of PEP carboxykinase?
It is a decarboxylation followe by a phosphoryl transfer (from GTP)
What cofactor is required for PEP carboxykinase to function?
GTP
What does GTP stand for?
Guanosine triphosphate
What is the structure of GTP?
What type of enzyme catalyzes the conversion of oxaloacetate to malate?
Dehydrogenase
What cofactor is required for the dehydrogenation of oxaloacetate to malate?
NADH
The conversion of oxaloacetate to malate operates under what type of mechanism?
Redox
What is the mechanism for the interconversion of oxaloacetate and malate?
Add picture
Identify this compound.
Oxaloacetate
Identify this compound.
“Malate”
What is the mechanism for the conversion of oxaloacetate to phosphoenolpyruvate?
The first bypass reaction of gluconeogenesis is the conversion of pyruvate to oxaloacetate to phosphoenolpyruvate via the oxaloacetate-malate shuttle system. What is the second bypass reaction of gluconeogenesis?
The hydrolysis of fructose-1,6-bisphosphate to fructose-6-phosphate and inorganic phosphate
What is the mechanism for the conversion of fructose-1,6-bisphosphate to fructose-6-phosphate in gluconeogenesis?
Phosphoester hydrolysis
What is the name of the enzyme that catalyzes the hydrolysis of fructose-1,6-bisphosphate to fructose-6-phosphate in gluconeogenesis?
Fructose-1,6-bisphosphatase
What is the third bypass reaction of gluconeogenesis?
The conversion of glucose-6-phosphate to glucose via glucose-6-phosphatase
What is the mechanism of action for glucose-6-phosphatase?
Phosphoester hydrolysis
What are the last two reactions of gluconeogenesis (catalyzed by fructose-1,6-bisphosphatase and glucose-6-phosphatase) not ATP-producing reactions?
Because the reactants are not high-energy compounds
What is the energetic cost of gluconeogenesis?
Total: 6 ATP
All result from the conversion of pyruvate to oxaloacetate
What is the first mechanisnistic step of pyruvate carboxylase?
The activation of carbon dioxide from bicarbonate by enzymatic addition of a phosphate group
What does the “business end” of biotin facilitate in the pyruvate carboxylase mechanism?
The transfer of carbon dioxide from one active site to another via covalent bonding
Why is a phosphate group attached to carbon dioxide in the pyruvate carboxylase mechanism?
Because it is functions as an appropiate leaving group, enabling biotin to “capture” the carbon dioxide
Why is ATP needed in the pyruvate carboxylase mechanism?
As a source of phosphate
What is the carrier of carbon dioxide in the pyruvate carboxylase mechanism?
Biotin
Which molecule ultimately abstracts carbon dioxide from the carboxybiotin intermediate?
Pyruvate (C3 carbanion)
What is the intermediate named in pyruvate carboxylase called?
Carboxybiotin
Does pyruvate carboxylase facilitate the addition or removal of a carbon dioxide molecule?
The addition (to pyruvate)
What chemical changes does pyruvate carboxylase catalyze? In other words, what happens to pyruvate?
A carbon dioxide molecule (COO-) is added to pyruvate, resulting in a four carbon molecule with an additional terminal COO- group
What does malate dehydrogenase do to oxaloacetate?
Reduces the carbonyl to a hydroxyl group
Does PEP carboxykinase facilitate a carboxylation or decarboxylation?
Decarboxylation
What is the first step in the PEP carboxykinase mechanism?
Removal of CO2
What is the second step in the PEP carboxykinase mechanism?
Phosphoryl transfer from GTP
PEP carboxykinase catalyzes the conversion of oxaloacetate to what intermediate of glycolyis?
Phosphoenolpyruvate
Describe the mechanism of PEP carboxykinase?
The oxygen anion on C4 kicks back and expels CO2
A carbanion forms on C3, which then kicks back, forming an alkene with C2, and the carbonyl oxygen becomes phosphorylated
What is the second step in the pyruvate carboxylase mechanism?
A nitrogen atom in biotin is deprotonated and becomes a nucleophile, attacking the carbonyl in the activated CO2 molecule and expelling Pi
What is the third step in the pyruvate carboxylase mechanism?
After bicarbonate has been phosphorylated and dephosphorylated, forming the carboxybiotin intermediate, C3 of pyruvate is deprotonated; the resulting carbanion attacks the carbonyl of the carboxybiotin, ultimately transfering the carboxyl group to pyruvate and restoring biotin’s original configuration
