7. lipid metabolism

LIPID METAMOLISM

1. Unit-7 Metabolism of Lipids

2. Chapter contents Digestion & Absorption Bile acid & Fat Digestion Fat soluble Vitamins Transportation & Circulation Fatty acid Oxidation & Generation of Energy Deposition of Fat ( synthesis of Triacyl glycerol) Mobilization of Fat ( Catabolism of triacyl glycerol) Phospholipid Metabolism Lipid Transport Lipoproteins ( Chylomicrons,VLDL,LDL,HDL ) Clinical Correlates – a)Ketosis ;b) Steatorrhoea & malabsorption syndromes ; c)Fatty Liver ; d)Disorders of Plasma lipoprotein ( hypercholestrolemia & atheroscelerosis )

3. Digestion and Absorption of Lipids

4. A. PREPARATORY PHASE Digestion in mouth and stomach: It was believed earlier that little or no fat digestion takes place in the mouth . Recently a lipase has been detected called lingual lipase which is secreted by the dorsal surface of the tongue (Ebner’s gland ). Lingual lipase : Lingual lipase activity is continued in the stomach also where the pH value is low. Due to retention of food bolus for 2 to 3 hours, about 30 per cent of dietary triacyl glycerol (TG) may be digested . Lingual lipase is more active on TG having shorter FA chains and is found to be more specific for ester linkage at 3-position rather than position-1.

5. Gastric lipase: There is evidence of presence of small amounts of gastric lipase in gastric secretion. The overall digestion of fats, brought about by gastric lipase is negligible because: No emulsification of fats takes place in stomach The enzyme secreted in small quantity pH of gastric juice is not conducive which is highly acidic , whereas gastric lipase activity is more effective at relatively alkaline pH.

6. 2. Digestion in small intestine: The major site of fat digestion is the small intestine. This is due to the presence of a powerful lipase (steapsin) in the pancreatic juice and presence of bile salts, which acts as an effective emulsifying agent for fats . Pancreatic juice and bile enter the upper small intestine, the duodenum , by way of the pancreatic and bile ducts respectively . Secretion of pancreatic juice is stimulated by the: Passage of an acid gastric contents (acid chyme) into the duodenum By secretion of the GI hormones, secretin and CCKPZ.

7. – Secretin: Increases the secretion of electrolytes and fluid components of pancreatic juice . – Pancreozymin of CCK-PZ: Stimulates the secretion of the pancreatic enzymes. – Cholecystokinin of CCK-PZ: Causes contraction of the gallbladder and discharge the bile into the duodenum . Discharge of bile is also stimulated by secretin and bile salts themselves. – Hepatocrinin: Released by the intestinal mucosa stimulates more bile formation which is relatively poor in the bile salt content.

8. Pancreatic juice has been shown to contain a number of lipolytic enzymes: 1. Pancreatic lipase (steapsin) 2. Phospholipase A 2 (Lecithinase) 3. Cholesterol esterase. The pancreatic lipase is the most important which hydrolyses TG containing short-chain FA as well as long chain FA . Other two enzymes are required for phospholipids and cholesterol respectively.

9. Pancreatic lipase (steapsin ): 1. Role of bile salts in pancreatic lipase activity: Bile salts are required for proper functioning of the enzyme . (a) Bile salts help in combination of ‘lipase’ with two molecules of a small protein called as colipase in the intestinal lumen. This combination of lipase with colipase has two effects: Enhances the lipase activity of the intestinal pH. Also protects the enzyme against inhibitory effects of bile salts and against surface denaturation. (b) Bile salts also help in emulsification of fats .

10. The complete hydrolysis of fats (TG) produces glycerol and FA. Pancreatic lipase is virtually specific for the hydrolysis of primary ester linkage. It cannot readily hydrolyze the ester linkage of position 2. Digestion of TG molecule by pancreatic lipase proceeds First by removal of a terminal FA to produce diglyceride The other terminal FA is then removed to produce monoglyceride.

11. B. TRANSPORT PHASE Products of fat digestion, FFA and monoglycerides mainly enter the microvilli and the apical pole of the absorptive epithelial cells by simple diffusion. Short and medium chain FA and unsaturated FA are absorbed more readily than long chain FA. Products of digestion is taken up by SER and resynthesized into TG again by the enzymes in the intestinal epithelial cells. This maintains a sharp gradient of concentration within the mucosal cells that favours continued rapid diffusion of FA into the cell from intestinal lumen. There is a merging of SER into RER- in which probably enzymes for TG resynthesis are formed as well as the protein component (apo-B 48 ) of lipoprotein complex, chylomicron .  

12. C. TRANSPORTATION PHASE

13. Resynthesized TG cannot pass to lymphatics nor to portal blood as it is insoluble. Hence, converted to lipoprotein complex chylomicrons. Each droplet of hydrophobic and water insoluble TG gets covered with a layer of hydrophilic PL, cholesterol/cholesterol esters and an apoprotein apo-B48.

14. Fat soluble vitamins Found in the fats and oils of food. Absorbed into the lymph and carried in blood with protein transporters = chylomicrons. Stored in liver and body fat and can become toxic if large amounts are consumed.

15. The fat-soluble vitamins, A, D, E, and K, are stored in the body for long periods of time and generally pose a greater risk for toxicity when consumed in excess than water-soluble vitamins. Eating a normal, well-balanced diet will not lead to toxicity in otherwise healthy individuals

16. Unlike water-soluble vitamins, these vitamins dissolve in fat and are stored in body tissues. Because they are stored, over time they can accumulate to dangerous levels and can lead to a condition called hypervitaminosis, meaning excess amounts of a vitamin in the body, if more than the recommended amount is taken.

17. This substance, which is produced in the liver, flows into the small intestine, where it breaks down fats. Nutrients are then absorbed through the wall of the small intestine. Upon absorption, the fat-soluble vitamins enter the lymph vessels before making their way into the bloodstream.

18. Transporting fat through water based blood and lymphatic system is a challenge. Short and medium chain fatty acids travel via cardiovascular system to liver. Long chain enter lymph system. Fat is transported via lipoproteins Lipid core, shell composed of protein , phospholipid & cholesterol that allow transportation.

19. Degradation of Fatty Acids Fatty acids are aerobically oxidized by β -oxidation in the mitochondrial matrix of lever, muscle, heart, renal cortex and brown adipose tissue for energy production during starvation, carbohydrate deprivation and sustained muscular work. β -Oxidation. β -oxidation takes place in mitochondrion. Several enzymes known collectively as FA-oxidase system are found in the mitochondrial matrix. These enzymes catalyze the oxidation of fatty acid to acetyl-CoA.

20. Activation of Fatty Acid: Fatty acids are in cytosol of the cell. Fatty acids must be first activated so that they participate in metabolic pathway. The activation requires energy, which is provided by ATP. In presence of ATP and coenzyme A, the enzyme acyl-CoA synthetase (thiokinase) catalyzes the conversion of a free fatty acid to an active fatty acid (Acyl-CoA). The acyl-CoA synthetase enzyme is found in the endoplasmic reticulum and inside and outside of mitochondria. Active fatty acid (acyl-CoA) are formed in the cytosol, where as β –oxidation of fatty acid occurs in mitochondrial matrix. Acyl-CoA is impermeable to mitochondrial membrane. Long chain activated fatty acid penetrate the inner mitochondrial membrane only in combination with carnitine ( β –hydroxy ϒ - trimethyl ammonium butyrate). It is considered as a carrier molecule.

21. Carnitine next carries the long chain fatty acyl group across the inner mitochondrial membrane to the mitochondrial matrix. Acyl-CoA molecule diffuse through the outer mitochondrial membrane to enter the outer mitochondrial compartment where carnitine palmitoyl transferase I of the outer mitochondrial membrane transfers acyl group of acyl-CoA to carnitine, forming acylcarnitine. This is transported inward across the inner membrane by an inner protein, carnitine- acylcarnitine translocase, in exchange of free carnitine brought out from the matrix (Carnitine- acylcarnitine antiport). Carnitine palmitoyl transferase II on the matrix surface of the inner membrane then transfers the acyl group of acyl carnitine to coenzyme A of the matrix, producing acyl-CoA and free carnitine.

22. β – Oxidation reaction. Four β –Oxidation reactions are repeated cyclically in the mitochondrial matrix, each cycle shortening the fatty acid chain of acyl-CoA by releasing two carboxy terminal carbons of the acyl chain as acetyl-CoA.

23. This impedes entry of acetyl-CoA into Krebs cycle. Acetyl-CoA in liver mitochondria is converted then to ketone bodies , acetoacetate & b -hydroxybutyrate. During fasting or carbohydrate starvation, oxaloacetate is depleted in liver due to gluconeogenesis.

24. KETONE BODIES: AN ALTERNATE FUEL FOR CELLS Liver mitochondria have the capacity to convert acetyl CoA derived from fatty acid oxidation into ketone bodies. The compounds categorized as ketone bodies are acetoacetate, 3-hydroxybutyrate (also called β-hydroxybutyrate), and acetone. Acetoacetate and 3-hydroxybutyrate are transported in the blood to the peripheral tissues. There they can be reconverted to acetyl CoA, which can be oxidized by the TCA cycle. Ketone bodies are important sources of energy for the peripheral tissues because   1) They are soluble in aqueous solution and, therefore, do not need to be incorporated into lipoproteins or carried by albumin as do the other lipids. 2 ) They are produced in the liver during periods when the amount of acetyl CoA present exceeds the oxidative capacity of the liver; and 3) They are used in proportion to their concentration in the blood by extrahepatic tissues, such as the skeletal and cardiac muscle and renal cortex. Even the brain can use ketone bodies to help meet its energy needs if the blood levels raise sufficiently; thus, ketone bodies spare glucose. This is particularly important during prolonged periods of fasting.

25. Disorders of fatty acid oxidation present with the general picture of hypoketosis (due to decreased availability of acetyl CoA) and hypoglycemia (due to increased reliance on glucose for energy). During a fast, the liver is flooded with fatty acids mobilized from adipose tissue. The resulting elevated hepatic acetyl CoA produced primarily by fatty acid degradation inhibits pyruvate dehydrogenase and activates pyruvate carboxylase. The OAA thus produced is used by the liver for gluconeogenesis rather than for the TCA cycle. Therefore, acetyl CoA is channelled into ketone body synthesis. This pushes acetyl CoA away from gluconeogenesis and into ketogenesis.

26. Use of ketone bodies by the peripheral tissues: ketolysis Although the liver constantly synthesizes low levels of ketone bodies, their production becomes much more significant during fasting when ketone bodies are needed to provide energy to the peripheral tissues. 3- Hydroxybutyrate is oxidized to acetoacetate by 3-hydroxy butyrate dehydrogenase, producing NADH (Figure 16.23). Aceto acetate is then provided with a CoA molecule taken from succinyl CoA by succinyl CoA:acetoacetate CoA transferase (thiophorase). This reaction is reversible, but the product, acetoacetyl CoA, is actively removed by its conversion to two acetyl CoA. Extrahepatic tissues, including the brain but excluding cells lacking mitochondria (for example, red blood cells), efficiently oxidize acetoacetate and 3-hydroxybutyrate in this manner. In contrast, although the liver actively produces ketone bodies, it lacks thiophorase and, therefore, is unable to use ketone bodies as fuel.

27. Triacylglycerol synthesis and degradation Biosynthesis of Triacylglycerols Triacylglycerol is the principle lipid or fat depots in the body and also constitute the major dietary lipids. These are synthesized mostly by microsomal enzymes of hepatocytes, adipocytes, mammocytes, enterocytes and muscle fibres in the following ways:- Glycerophosphate pathway Substrates required for the synthesis of Triacylglycerol are: ( a) glycerol-3 phosphate (b) Acyl-CoA 1 . Fatty acid thiokinases use ATP and Co-A to thioesterify fatty acids to acyl-CoA molecules. 2. Glycerol 3-phosphate is produced in two alternative ways. (i) In hepatocytes and mammocytes, glycerol kinase uses ATP to phosphorylate glycerol. (ii) In adipocytes, enterocytes and skeletal muscles, glycerol 3-phosphate dehydrogenase uses NADH to reduce dihydroxyacetone phosphate from glycolysis to glycerol 3-phosphate.

28. 3. Glycerol 3-phosphate acyltransferase transfers an acyl group, mostly from a saturated acyl-CoA to glycerol 3-phosphate to change the latter to lysophosphatidic acid. 4. Lysophosphatidate acyltransferase transfers an acyl group from an acyl-CoA to lysophosphatidic acid, changing the latter to phosphatidic acid. 5. Phosphatidate phosphohydrolase hydrolyzes the phosphoester bond in phosphatidic acid to release Pi and 1,2-diacylglycerol. 6. Diacylglycerol acyltransferase transfers an acyl group, mostly from acyl-CoA to 1,2 diacylglycerol to produce Triacylglycerol.

29.   Intestinal monoacylglycerol shunt In enterocytes, (a) monoacylglycerol acyltransferase transfers an acyl group, from an acyl-CoA to 2-monoacylglycerol produced by fat digestion, to change it to 1,2 diacylglycerol. ( b) diacylglycerol acyltransferase next transfers another acyl group from an acyl-CoA to 1,2-diacylglycerol to produce Triacylglycerol.

30. Steatorrhoea and malabsorption syndrome Normally more than 95% of ingested lipid is absorbed. When a large fraction is excreted in the feces , it is called steatorrhea. Measurement of fecal lipid with adequate lipid intake is a sensitive indicator of lipid malabsorption. Malabsorption can result from impairment in lipolysis (Table 12-6), micelle formation (Table 12-7), absorption, chylomicron formation, or transport of chylomicrons via the lymph to blood.

31. Fatty Liver TG and FA synthesized or taken up by the liver, are rapidly incorporated into lipoproteins and then mobilized to extrahepatic tissues through the plasma mainly as VLDL. However, under the following conditions mentioned below, the liver gets loaded with too much lipids, leading to fibrosis of hepatocytes, cirrhosis and hepatic failure. The amount of lipids in the liver at any given time is the resultant of several influences, some acting in conjunction with and some in opposition to other. Normal liver contains about 4% as total lipids, three fourths of which is phospholipids and one-fourth as neutral fats

32. a) Factors that tend to increase the fat content of liver are: Influx of dietary lipids. Plasma free fatty acids may rise due to a high- fat diet. The liver then takes up far more fatty acids and produces far more TG from them than what it can incorporate into lipoproteins like VLDL for mobilization to extrahepatic tissue tissues. This may accumulate too much fat in the liver. Synthesis of FA from carbohydrates and proteins. Plasma free fatty acids may rise due to an increased adipose tissue lipolysis during starvation and diabetes. Mobilization of FA from depot to liver. Administration of puromycin or poisoning with CCl4, CHCl3, lead, phosphorus or arsenic may cause fatty liver by blocking the synthesis of apolipoprotein B-100 in the liver and consequently reducing VLDL synthesis.

33. Hypercholesterolemia Familial hypercholesterolemia (FH) is an inborn error of metabolism due to a defective LDL-receptor protein. Five classes of mutations have been identified consisting of more than 150 different alleles. The defects in the receptor function can be grouped into five types: 1. The receptor may not be synthesized at all; 2. The receptor may not be transported to the surface; 3. The receptor may fail to bind LDL; 4. The receptor may fail to cluster in coated pits; or 5. The receptor may fail to release LDL in the endosome.

34. ELECTRON TRANSPORT CHAIN Energy-rich molecules, such as glucose, are metabolized by a series of oxidation reactions ultimately yielding CO 2 and water. The metabolic intermediates of these reactions donate electrons to specific coenzymes—nicotinamide adenine dinucleotide (NAD + ) and flavin adenine dinucleotide (FAD)—to form the energy-rich reduced coenzymes, NADH and FADH 2 .

35. These reduced coenzymes can, in turn, each donate a pair of electrons to a specialized set of electron carriers, collectively called the electron transport chain. As electrons are passed down the electron transport chain, they lose much of their free energy. Part of this energy can be captured and stored by the production of ATP from ADP and inorganic phosphate (Pi). This process is called oxidative phosphorylation. The remainder of the free energy not trapped as ATP is used to drive ancillary reactions such as Ca 2+ transport into mitochondria, and to generate heat.

36. A. Mitochondrion The electron transport chain is present in the inner mitochondrial membrane and is the final common pathway by which electrons derived from different fuels of the body flow to oxygen. Electron transport and ATP synthesis by oxidative phosphorylation proceed continuously in all tissues that contain mitochondria. 1. Membranes of the mitochondrion: The components of the electron transport chain are located in the inner membrane. Although the outer membrane contains special pores, making it freely permeable to most ions and small molecules, the inner mitochondrial membrane is a specialized structure that is impermeable to most small ions, including H + , Na + , and K + , and small molecules such as ATP, ADP, pyruvate, and other metabolites important to mitochondrial function. Specialized carriers or transport systems are required to move ions or molecules across this membrane. The inner mitochondrial membrane is unusually rich in protein, half of which is directly involved in electron transport and oxidative phosphorylation. The inner mitochondrial membrane is highly convoluted. The convolutions, called cristae, serve to greatly increase the surface area of the membrane. 2. Matrix of the mitochondrion: This gel-like solution in the interior of mitochondria is 50% protein. These molecules include the enzymes responsible for the oxidation of pyruvate, amino acids, fatty acids (by beta-oxidation ), and those of the tricarboxylic acid (TCA) cycle. The synthesis of glucose, urea, and heme occur partially in the matrix of mitochondria. In addition, the matrix contains NAD + and FAD (the oxidized forms of the two coenzymes that are required as hydrogen acceptors) and ADP and Pi, which are used to produce ATP .

37. B. Organization of the electron transport chain The inner mitochondrial membrane can be disrupted into five separate protein complexes, called Complexes I, II, III, IV, and V . Complexes I–IV each contains part of the electron transport chain Each complex accepts or donates electrons to relatively mobile electron carriers, such as coenzyme Q and cytochrome c . Each carrier in the electron transport chain can receive electrons from an electron donor, and can subsequently donate electrons to the next carrier in the chain. The electrons ultimately combine with oxygen and protons to form water . This requirement for oxygen makes the electron transport process the respiratory chain, which accounts for the greatest portion of the body’s use of oxygen. Complex V is a protein complex that contains a domain ( Fo ) that spans the inner mitochondrial membrane, and a domain (F1) that appears as a sphere that protrudes into the mitochondrial matrix. Complex V catalyzes ATP synthesis and so is referred to as ATP synthase. C. Reactions of the electron transport chain With the exception of coenzyme Q, all members of this chain are proteins. These may function as enzymes as is the case with the dehydrogenases, may contain iron as part of an iron–sulphur center, may be coordinated with a porphyrin ring as in the cytochromes, or may contain copper as does the cytochrome a + a3 complex.

38. 1. Formation of NADH: NAD+ is reduced to NADH by dehydrogenases that remove two hydrogen atoms from their substrate. Both electrons but only one proton (that is, a hydride ion, :H–) are transferred to the NAD+, forming NADH plus a free proton, H+. 2. NADH dehydrogenase: The free proton plus the hydride ion carried by NADH are next transferred to NADH dehydrogenase, a protein complex (Complex I) embedded in the inner mitochondrial membrane. Complex- II has a tightly bound molecule of flavin mono nucleotide (FMN, a coenzyme structurally related to FAD, that accepts the two hydrogen atoms (2e– + 2H+), becoming FMNH2. NADH dehydrogenase also contains iron atoms paired with sulfur atoms to make iron– sulfur centers. These are necessary for the transfer of the hydrogen atoms to the next member of the chain, coenzyme Q (ubiquinone). 3. Coenzyme Q: Coenzyme Q (CoQ) is a quinone derivative with a long, hydrophobic isoprenoid tail. It is also called ubiquinone because it is ubiquitous in biologic systems. CoQ is a mobile carrier and can accept hydrogen atoms both from FMNH2, produced on NADH dehydrogenase (Complex I), and from FADH2, produced on succinate dehydrogenase (Complex II), Glycerophosphate dehydrogenase and acyl CoA dehydrogenase CoQ transfers electrons to Complex III. CoQ, then, links the flavoproteins to the cytochromes.