Glycolysis and TCA cycle Flashcards
List the three key steps in glycolysis
1 - Activation of glucose to Glucose-6-phosphate (G6P)
2 - Second phosphorylation to Fructose-1,6-bisphosphate
3 - Synthesis of 2nd ATP by 2nd substrate level phosphorylation
What is the process and enzymes involved in the activation of glucose?
Goal of reaction: D-Glucose becomes G-6-P
Enzymes: Hexokinase or glucokinase
Irreversible reaction
G-6-P: Product of reaction inhibits hexokinase
Process: Phosphorylation gives glucose a negative charge, locking it in the cell, also conserves the energy of breaking the ATP bond by transferring it to G-6-P, lowers the activation energy of the next step by more easily binding to the enzyme.
What is the process and enzymes involved in the second phosphorylation step of glycolysis?
Goal of reaction: Fructose-6-phosphate converts to Fructose-1,6-phosphate
Enzymes: Phosphofructokinase-1
RATE LIMITING STEP! Major point of control of flow of glucose through glycolysis. Irreversible reaction. Sets up compound for cleavage step that releases energy.
What is the process and enzymes involved in the synthesis of ATP from glycolysis?
Goal of reaction: 2 phosphoenol pyruvate + 2 ADP -> 2 pyruvate + 2 ATP, results in the only net ATP gain of glycolysis.
Enzyme: Pyruvate kinase, deficiency of this enzyme is 2nd most common cause of enzyme linked hemolytic anemia (no glycolysis for RBCs)
Pyruvate now available for TCA cycle in presence of oxygen or lactate in absence of oxygen
Describe the enzymes responsible for initial phosphorylation of glucose and their regulation
Hexokinase: Non-specific Present in all cells Low Km for sugars Inhibited by G-6-P Tightly regulated enzyme
Glucokinase:
Selective only for glucose
Present ONLY in pancreatic beta cells and liver
High Km for glucose
Inhibited by F-6-P
The efficient glucose transporter in hepatocytes maintains a cytosolic Glu concentration close to that of blood. Glucokinase allows the liver to regulate blood glucose by transporting it into the liver and entering glycolysis.
Describe the enzyme responsible for the rate limiting step of glycolysis and its regulation
Phosphofructokinase 1:
Allosteric enzyme sensitive to levels of ATP (ATP/citrate inhibits, AMP induces)
Potently activated by F-2,6-BP, created by PFK2
PFK2 is a bifunctional Kinase & Phosphatase and can create F-6-P -> F-2,6-BP -> F-6-P
F-2,6-BP also inhibits gluconeogenesis (while stimulating glycolysis)
Fed state = Increased F-6-P -> F-2,6-P via PFK2 -> Increased PFK1 activity -> Increased glycolysis
Fasted state = insulin drops, PFK2 is phosphorylated and switches to F-2,6-bisphosphatASE -> reduced F-2,6-BP -> reduced PFK1 activity and increased gluconeogensis
Describe the enzyme responsible for the final creation of pyruvate and its regulation
Pyruvate kinase:
2nd substrate level phosphorylation, stimulated by F-1,6-BP in glycolysis.
Inhibited by ATP, alanine and PKA
Promotes gluconeogenesis when sufficient cellular energy exists. During fasting, glucagon-dependent inactivation of liver pyruvate kinase through PKA phosphorylation prevents glycolysis and assures glucose synthesis. Enzyme is reactivated by dephosphorylation.
Lack of this enzyme is pyruvate kinase deficiency (hemolytic anemia).
Describe the situation in which flux through glycolysis is increased
High insulin:glucagon ratio means lots of glucose available for cell (or brain) to eat.
This ratio causes decreased cAMP and decreased PKA activity.
Decreased PKA activity favors dephosphorylation of PFK2 complex, leading to increased F-2,6-BP -> increased PFK1 activity and increased glycolysis.
Describe the situation in which flux through glycolysis is decreased
Low Insulin:Glucagon ratio means little glucose available for cells (or brain) to eat.
Glucagon stimulates cAMP creation leading to increased PKA activity.
Increased PKA favors phosphorylated PFK2 which undergoes conformational change to F-2,6-bisphosphatASE and dephosphorylates F-2,6-BP to F-6-P. Loss of F-2,6-BP inhibits glycolysis and stimulates gluconeogensis via (the now dis-inhibited) F-1,6-bisphosphatASE.
What is the final product of anaerobic glycolysis?
Pyruvate is converted to lactate in order to regenerate NAD from the NADH created in glycolysis. This is known as lactic acid fermentation, and occurs in RBC, sperm, and retinal cells. In the liver and the heart the NADH:NAD ratio is lower than in exercising muscle, so they tend to convert lactate into pyruvate. Lactic acidosis is very damaging to cardiac tissue.
What is the final product of aerobic glycolysis?
In aerobic glycolysis, the NADH generated is oxidized by the electron transport chain to produce NAD and 2 ATP. Pyruvate is converted into Acetyl-CoA and generates much more energy via the TCA cycle/Electron Transport Chain.
List the substrates that provide carbon skeletons to the TCA cycle
Carbon skeletons enter the TCA cycle mostly as Acetyl-CoA, though there are a few differences
Sugars via pyruvate -> Acetyl-CoA
Fats via beat oxidation -> Acetyl-CoA
Amino acids enter at several points as a-ketoglutarate, succinate, and alanine enters as Acetyl-CoA
What are the key intermediates of the TCA cycle?
Acetyl-CoA Citrate a-ketoglutarate Succinyl-CoA/Succinate Fumarate Oxaloacetate
Why is Acetyl-CoA important in the TCA?
Acetyl-CoA is important because it serves as a “node” where many metabolic pathways connect (carbs, fats, one AA). There must be an input of 2 carbon Acetyl group to start the TCA cycle, both carbons are lost in the TCA cycle.
Why is citrate important in the TCA cycle?
Citrate is a noteworthy intermediate because citrate is an inhibitor of PFK1 in glycolysis. High citrate causes high flux through the TCA cycle. Citrate may also leave the TCA cycle to form fatty acids in de novo lipogenesis.
