Gluconeogenesis

1. Gluconeogenesis Dr.S.Chakravarty MD

2. Maintenance of blood glucose during various states

3. Gluconeogenesis is the process of synthesizing glucose or glycogen from non-carbohydrate precursors.

4. Biomedical Importance • Gluconeogenesis meets the needs of the body for glucose when insufficient carbohydrate is available from the diet or glycogen reserves. • A supply of glucose is necessary especially for the nervous system and erythrocytes. • Glucose is also important in maintaining the level of intermediates of the citric acid cycle even when fatty acids are the main source of acetyl-CoA in the tissues. • Gluconeogenesis clears lactate produced by muscle and erythrocytes and glycerol produced by adipose tissue. • In ruminants, propionate is a product of rumen metabolism of carbohydrates, and is a major substrate for gluconeogenesis. • Failure of gluconeogenesis is usually fatal. Hypoglycemia causes brain dysfunction, which can lead to coma and death.

5. Tissues :- Liverand Kidney are the major gluconeogenic tissues, but the small intestine may also be a source of glucose in the fasting state. • Subcellular site :- Partly mitochondrial and partly cytosolic.

6. Substrates for Gluconeogenesis Glucogenic amino acids Lactate Glycerol Propionate

7. Glucogenic amino acids α-ketoglutarate Glutamate AlaninePyruvate • α-ketoglutarate Glutamate • AspartateOxaloacetate Alaninetransaminase (ALT) Aspartatetransaminase (ALT)

8. Entry points of Glucogenic amino acids

9. Lactate NAD+ NADH + H+ Lactate Pyruvate Lactate Dehydrogenase (LDH)

11. Glycerol ATP ADP Glycerol Glycerol 3 –P This enzyme is absent in adipose tissue . Glycerol kinase

12. Conversion of Glycerol to Glucose: Triglycerides FASTING OR LOW GLUCOSE Glycerol Fatty acids  Beta oxidation Glycerol kinase Liver Acetyl Co-A Glycerol 3- PO4 NAD+ Glycerol-3-po4 dehydrogenase NADH Dihydroxyacetone phosphate

13. Propionate – From oxidation of odd chain fatty acids

14. The pathway • Thermodynamic Barriers Prevent a Simple Reversal of Glycolysis. • Three nonequilibrium reactions in glycolysis catalyzed by hexokinase, phosphofructokinaseandpyruvatekinase, prevent simple reversal of glycolysis for glucose synthesis.

15. Step 1: Pyruvate Energy derived from fatty acid oxidation ATP Pyruvatecarboxylase CO2 ADP ABC carboxylase (Mitochondria) Oxaloacetate GTP GTP derived from succinatethoikinase Phosphoenolpyruvatecarboxykinase CO2 GDP USMLE CONCEPT!!! (cytosol) Phosphoenolpyruvate Problem --Mitochondrial membrane is impermeable to OAA!!

16. Oxaloacetate formed in the mitochondria enters cytosol through Malate Oxaloacetate NAD NADH Malate dehydrogenase Malate Mitochondria cytosol Malate NADH Malate dehydrogenase NAD Oxaloacetate

17. The next few steps are reversal of Glycolysis till Fructose 1,6 bisphosphate is formed.

18. Step 2: Conversion of fructose 1,6 bisphosphate to fructose 6-PO-4 Glucose ADP, AMP ATP Fructose -6-PO4 (+) (+) PFK -1 Fructose 1,6 bisphosphatase (+) (-) Fructose 1,6 Bisphosphate Fructose 2,6 Bisphosphate Fructose 2,6 Bisphosphate Pyruvate Glycolysis Gluconeogenesis

19. Step 3: Conversion to Glucose Glucose ATP PO4 Glucokinase Glucose-6-phosphatase ADP Glucose-6-po4 Glycolysis Gluconeogenesis

20. Glucose 6-Phosphate & Glucose & Glucose to glycogen(not shown) Fructose 1,6-Bisphosphate & Fructose 6-Phosphate 1.Pyruvate & Phosphoenolpyruvate GLUCONEOGENESIS

21. Regulation • Glycolysis and Gluconeogenesis are regulated reciprocally. Regulation Covalent modification Induction / Repression Allosteric

22. Induction & Repression of Key Enzymes Requires Several Hours.

23. Table 20.1 Harper 29th page 190

24. Covalent Modification by Reversible Phosphorylation Is Rapid Glucagon and epinephrine inhibit glycolysis and stimulate gluconeogenesis in the liver by increasing the concentration of cAMP. cAMPcAMP-dependent protein kinase phosphorylation and inactivation of pyruvatekinase. They also affect the concentration of fructose 2,6-bisphosphate which is the most potent positive allostericeffector of of Phosphofructokinase -1 and inhibitor of Fructose 1,6 bisphosphatase .

25. Insulin Fructose -6-po4 PFK-2 Glucagon PFK-1 Fructose -2,6- Bisphosphate Fructose -1,6- Bisphosphate USMLE concept!!! Regulation of PFK -1 :

26. Allosteric Regulation is Instantaneous !! Glucagon (+) Source of pyruvate ? Pyruvate PyruvateCarboxylase Pyruvate dehydrogenase (+) (-) Oxaloacetate Acetyl Co-A Acetyl Co-A is the allosteric activator of pyruvatecarboxylase = allosteric regulation What is the source of acetyl Co-A during starvation ?

27. Allosteric Modification Is Instantaneous • Acetyl -CoA as an is an allosteric activator of Pyruvatecarboxylase. Phosphofructokinase (phosphofructokinase-1 )is inhibited by citrate and by normal intracellular concentrations of ATP and is activated by 5'AMP. (5'AMP acts as an indicator of the energy status of the cell. ) • When ATP is used AMP increases sensitive signal for energy state of the cell.

28. Substrate cycles allow fine tuning , rapid response & generate heat ATP ADP A B Pi H20 NET FLUX OF B = 0 +heat 100 120 Other factors that favor AB 80 100 40 At rest the activity of PFK IS 10x greater than F1,6BPhosphatase . During Muscle contraction , PFK activity increases and F1,6BPhosphatase falls so that rate of Glycolysis becomes 1000 fold higher than resting state. Futile cycle may occur physiologically for generation of heat. It takes place to a great extent in animals undergoing arousal from hibernation, when body temp is much lower .

29. Clinical aspects 1. Pyruvatecarboxylase deficiency (A.R)- 1 in 25,000 births –characterized by Hypoglycemia , lactic acidosis and Mental retardation . 2. Fructose 1,6bisphosphatase deficiency – lactic acidosis and hypoglycemia .Treatment – feed high carb. Diets and avoidance of fasting . 3. Hypoglycemia during pregnancy and in neonates – Increased risk of maternal hypoglycemia if there are long intervals b/w meals . Premature babies are more susceptible to hypoglycemia. They have immmature non functional enzymes for gluconeogenesis and low adipose tissue mass.

30. Pyruvate carboxylase deficiency:

31. Malfunctioning of citric acid cycle – def of oxaloacetate • Malfunctioning of gluconeogenesis – def of oxaloacetate - Hypoglycemia • Malfunctioning of urea cycle – Def of Aspartate • Acetyl Co-A forms ketone bodies-KETOGENESIS

32. ALCOHOL DEHYDROGENASE • Alcohol  Acetaldehyde Acetate • Excess NADH  • EXCESS LACTATE from PYRUVATE • Excess Malate FROM OAA • Excess Glycerol 3 P from DHAP •  No or less Gluconeogenesis!!Hypoglycemia NAD NADH NAD NADH

33. Immediately after completing a 25-mile marathon race, a healthy 24-yr old man was extremely dehydrated and thirsty. He quickly consumed a 6-pack of ice-cold beer and shortly thereafter became very weak and light-headed and nearly fainted. He complained of muscle cramping and pain. What is the most probable cause ? • Excess lactate in blood • Excess Alcohol in blood • Excess NADH • Dehydration • Electrolyte imbalance

34. Which of the following enzymes can be induced genetically by hormones in a person with prolonged history of fasting? • Glucokinase • Pyruvate Carboxylase • PFK-1 • Acetyl co-A Carboxylase • Phosphofructokinase

35. In the citric acid cycle, succinatethiokinase catalyzes the cleavage of the succinyl –Co-A to succinate with formation of a high energy compound. This compound can then be used by the body in which of the following biochemical pathways? • Oxidative phosphorylation • Gluconeogenesis • Formation of creatine phosphate • Cholesterol synthesis • Fatty acid synthesis

36. Thank you