Fatty acid oxidation Ketone bodies Fatty acid synthesis

1. AV. Lipid metabolism 1-2. 2011 Fatty acid oxidationKetone bodiesFatty acid synthesis

2. Triacylglycerols Major energy reserve Oxidation: 9 kcal/g (for carbohydrates: 4 kcal/g) 11 kg of 70 kg total body weight Site of accumulation:cytoplasm of ADIPOSECELLS Adipose tissue -specialized for synthesis, storage, mobilization of lipids Lipases

5. The most important fatty acids

6. Mobilization of triacylglycerols stored in adipose tissue Glukagon, adrenalin, ACTH ↓ receptor activation ↓ protein kinase A ↓ phosphorylation of Perilipin (when dephosphorylated it inhibits the access of lipases to TG and DG + phosphorylation of HSL (hormon-sensitive lipase) activation ↓ CGI dissociates from Perilipin then associates with ATGL (adipocyte trigliceride lipase) ↓ TG→DG + fatty acid ↓ HSL: DG→MG + fatty acid ↓ MGL (monoacylglycerol lipase): MG→G + fatty acid Lehninger, Principals of Biochemistry, 2013

7. ADIPOSE TISSUE TG Fatty acids CIRCULATION Fatty acids - bound to albumin (10 fatty acids/albumin monomer) (free fatty acids, FFA) MUSCLE, HEART MUSCLE, RENAL CORTEX Fatty acid activation, transport into the mitochondria, β-oxidation

8. Site of β-oxidation: mitochondria

9. Activation of fatty acids:Fatty acid + ATP + CoA <------> Acyl-CoA + PPi + AMPAcyl-CoA synthetase Fast PPi hydrolysis reaction is irreversible in vivo

10. Site of fatty acid activation: cytosolic side of the mitochondrial outer membrane

11. Triacylglycerols Major energy reserve Oxidation: 9 kcal/g (for carbohydrates: 4 kcal/g) 11 kg of 70 kg total body weight Site of accumulation:cytoplasm of ADIPOSECELLS Adipose tissue -specialized for synthesis, storage, mobilization of lipids Lipases

14. The most important fatty acids

15. Mobilization of fatty acids from the adipose tissue Glukagon, adrenalin, ACTH Receptor activation Protein kináz A Phosphorylation of Perilipin (when dephosphorylated it inhibits the access of lipase to TG, phosphorylated perilipin has no such effect) + Phosphorylation of Hormon-sensitive lipase (activation) Mobilization of fatty acids from TG Lipid droplet

16. ADIPOSE TISSUE TG Fatty acids CIRCULATION Fatty acids - bound to albumin (10 fatty acids/albumin monomer) (free fatty acids, FFA) MUSCLE, HEART MUSCLE, RENAL CORTEX Fatty acid activation, transport into the mitochondria, β-oxidation

17. Site of β-oxidation: mitochondria

18. Activation of fatty acids:Fatty acid + ATP + CoA <------> Acyl-CoA + PPi + AMPAcyl-CoA synthetase Fast PPi hydrolysis reaction is irreversible in vivo

19. Site of fatty acid activation: cytosolic side of the mitochondrial outer membrane

20. Transport of fatty acids into the mitochondria Transport of Long-chain (12-18 C atom) fatty acids with carnitine

21. Transport of fatty acids into the mitochondria carnitine-acyltransferase I (CPT I) Transport of carnitine-acylcarnitine is the rate-limiting step and most important control point in fatty acid oxidation carnitine-acyltransferase II (CPT II) Fatty acids with <12 C enter mitochondria without carnitine and are activated in the mitochondria

22. Three isoenzymes: -long-chain fatty acids (C 12-18) -medium chain fatty acids (MCAD, 4-14) -short-chain fatty acids (4-8) MCAD deficiency is relatively frequent

23. Specific for L-stereoisomere

24. β-oxidation of fatty acids Oxidation of fatty acids with >12 C is carried out by a multienzyme bound to the mitochondrial inner membrane, in which the last three enzymes are tightly associated (trifunctional protein), when the chain is < 12 C soluble enzymes in the matrix continue the oxidation

27. Conversion of glycerol to glycolysis intermediate – in the liver glycerol kinase Glycerol-P dehydrogenase

28. Regulation of fatty acid oxidation hormones (adrenaline, glucagon) High energy state NADH Acetyl-CoA Malonyl-CoA Inhibiton of Perilipin Activation of hormone-sensitive lipase thiolase Inhibition of 3-hydroxyacyl CoA dehydrogenase Inhibition of carnitine acyltransferase I Increased level of free fatty acids Entry of fatty acids into mitochondria is inhibited Inhibition of ß-oxidation Oxidation Oxidation

29. Long-term regulation: PPAR (peroxisome proliferator/activated receptors) nuclear receptor – transcription factors PPARα – muscle, adipose tissue, liver regulate the transcription of fatty acid transporters, CPT I és CPT II, and acyl-CoA dehydrogenase - energy need (fasting, between-meal periods) PPARα activation transcription of enzymes of fatty acid oxidation - fetus - principal fuels for heart: glucose and lactate -neonatal– fatty acid Regulation of metabolic transitionby PPARα - sustained exercise – PPARα expression in muscles

30. The most common genetic defect in fatty acid oxidation Acyl-CoA dehydrogenase deficiency For the medium length acyl-CoA dehydrogenase Prevalence: 1:40 – mutation in one of the chromosomes 1:10000 – two mutant copies – disease manifestation symptoms in the first years -hypoglycemia – with decreased ketone body formation (decreased fatty acid oxidation and gluconeogenesis in the liver) -accumulation of lipids in the liver -vomiting, drowsiness Therapy: frequent carbohydrate-rich meals + carnitine supply

31. Deficiency of carnitine transport into the mitochondria Long-chain fatty acid transport Carnitine deficiency -high-affinity plasma membrane transporter (heart, kidney, muscle – but not liver) muscle cramps – weakness – death addition of carnitine -secondary carnitine deficiency due to deficiency on β-oxidation acyl-carnitine in the urine Carnititne acyltransferase deficiency most common - CPT II gene mutation – partial loss of enzyme activity muscle weakness when more serious – hypoglycemia with decreased ketone body formation

32. FORMATION OF KETONE BODIES FATTY ACID GLUCOSE ß-oxidation GLUCONEOGENESIS ACETYL-CoA PYRUVATE ANAPLEROTIKUS REAKCIÓ OXALOACETATE Ketone bodies CITRATE Fatty acid oxidation +lack of oxaloacetate fasting untreated diabetes

34. Synthesis of ketone bodies in the liver

35. Ketone bodies as fuels

36. Oxidation of ketone bodies in the extrahepatic tissues heart muscle striatal muscle kidney brain

37. Ketone bodiescan be regarded as a transport form of acetyl groups Important sources of energy: heart muscle,renal cortex (preference to glucose, 1/3 of the energy) brain - glucose is the major fuel but in starvation and diabetes brain uses acetoacetate

38. Fasting Diabetes high level of ketone bodies in the blood KETOSIS Formation in the liver exceeds the use in the periphery. Level of ketone bodies after an overnight fast: ~0.05 mM 2 days starvation: 2 mM (40-fold increase!) 40 days: 7 mM Ketone bodies

40. Fatty acid synthesis - repeated cycles – in each cycle the chain is extended by two carbons – four steps in each cycle Enzyme: fatty acid synthase Seven active site for different reactions in separate domains of a single large polypeptide

41. Fatty acid synthesis –Lipogenesis-not a reversal of degradation

42. Fatty acid synthesis:*Liver*Adipose tissue*Lactating mammary gland

43. Committed step in fatty acid synthesis: formation of malonyl-CoA from acetyl-CoA acetyl-CoA carboxylase - prosthetic group: biotin

44. Acetyl-CoA carboxylase has three activities in a single polypeptide Biotin – covalently bound to Lys έ-amino group 1. transfer of carboxyl group to biotin ATP-dependent

45. move of activated CO2 from the biotin carboxylase region to the transcarboxylase active site

46. 2. transfer of the activated carboxil group from biotin to acetyl-CoA

47. Critical SH-groups carry the intermediates during the synthesis of fatty acids Acyl carrier protein β-ketoacyl-ACP synthase SH group is the site of ently of malonyl group during fatty acid synthesis

48. Fatty acid synthase Acetyl group from Acetyl-CoA is transferred to Cys-SH of β-ketoacyl-ACP synthase (KS) by MAT

49. Malonyl group is transferred to ACP-SH by MAT

50. Acetyl group (from Acetyl-CoA) is transferred to the malonyl group on ACP (methyl terminal) Acetoacetyl-ACP