Metabolism MSLSC2001C04 Course Instructor Dr. Gautam Kumar Dr. Gautam Kr. Dept. of Life Sc 1 Dr. Gautam Kr. Dept. of Life Sc 2 Lipids Metabolism • Lipids are the principal form of stored energy in most organisms and major constituents of cellular membranes. • Specialized lipids serve as pigments (retinal, carotene) cofactors (vitamin K), detergents (bile salts), transporters (dolichols), hormones (vitamin D derivatives, sex hormones), extracellular and intracellular messengers (eicosanoids, phosphatidylinositol derivatives), and anchors for membrane proteins (covalently attached fatty acids, prenyl groups, and phosphatidylinositol). • Water-insoluble products from water-soluble precursors such as Acetate. • During Fatty acids synthesis ATP as a source of metabolic energy and a reduced electron carrier (usually NADPH). Dr. Gautam Kr. Dept. of Life Sc 3 Lipid Metabolism • Fatty acid synthesis takes place in the compartment in which NADPH is available for reductive synthesis (i.e., where the [NADPH]/[NADP] ratio is high) Subcellular localization of lipid metabolism Dr. Gautam Kr. Dept. of Life Sc 4 Acetyl-CoA Shuttle • Shuttle for transfer of AcetylDr.- GautamCoA Kr.groups Dept. of Life from Sc mitochondria to the cytosol5 • The mitochondrial outer membrane is freely permeable to all these compounds. • Pyruvate derived from amino acid catabolism in the mitochondrial matrix, or from glucose by glycolysis in the cytosol, is converted to acetyl-CoA in the matrix. • Acetyl groups pass out of the mitochondrion as citrate; in the cytosol they are delivered as acetyl-CoA for fatty acid synthesis. • Oxaloacetate is reduced to malate, which returns to the mitochondrial matrix and is converted to oxaloacetate. • Cytosolic malate is oxidation by malic enzyme to generate cytosolic NADPH; the pyruvate produced returns to the mitochondrial matrix. Dr. Gautam Kr. Dept. of Life Sc 6 Biosynthesis of Fatty Acids and Eicosanoids • Biosynthesis requires the participation of a three-carbon intermediate, Malonyl-CoA, • Malonyl-CoA Is Formed from Acetyl-CoA and Bicarbonate • Formation of malonyl-CoA from acetyl-CoA is an irreversible process, catalyzed by acetyl-CoA carboxylase. Dr. Gautam Kr. Dept. of Life Sc 7 • The bacterial enzyme has three separate polypeptide subunits. • In animal cells, all three activities are part of a single multifunctional polypeptide. • Plant cells contain both types of acetyl-CoA carboxylase. Acetyl-CoA carboxylase Biotin carboxylase, which activates CO2 Transcarboxylase, which transfers activated CO2 from biotin to acetyl-CoA, producing malonyl-CoA. Transcarboxylase Dr. Gautam Kr. Dept. of Life Sc 8 Biotinyl group - • The carboxyl group, derived from bicarbonate (HCO 3 ), is first transferred to biotin in an ATP-dependent reaction. • The biotinyl group serves as a temporary carrier of CO2, transferring it to acetyl-CoA in the second step to yield malonyl-CoA. Dr. Gautam Kr. Dept. of Life Sc 9 Carboxylation Dr. Gautam Kr. Dept. of Life Sc 10 Proteins of the Fatty Acid Synthase Complex of E. coli • Multienzyme complex, Fatty Acid Synthase • Structure differ in prokaryotes such as E. coli and in Eukaryotes • The Fatty Acid Synthase Complex has Seven ACP different active sites in E coli and some plants • Acyl carrier protein (ACP) of E. coli is a small protein containing the prosthetic group 4-phosphopantetheine Dr. Gautam Kr. Dept. of Life Sc 11 • Acyl carrier protein (ACP): The prosthetic group is 4- phosphopantetheine, which is covalently attached to the hydroxyl group of a Ser residue in ACP. • Phosphopantetheine contains the B vitamin pantothenic acid, also found in the coenzyme A molecule. • Its --SH group is the site of entry of malonyl groups during fatty acid synthesis. Malonyl transferase: Formation of malonyl -ACP Dr. Gautam Kr. Dept. of Life Sc 12 Fatty Acid Synthesis Proceeds in a Repeating Reaction Sequence The long carbon chains of fatty acids are assembled in a repeating four-step sequence 1.Condensation 2.Reduction Four-step process of fatty acid synthesis is the 3.Dehydration same in all organisms 4.Reduction Dr. Gautam Kr. Dept. of Life Sc 13 Fatty acid synthase ACP Dr. Gautam Kr. Dept. of Life Sc 14 Dr. Gautam Kr. Dept. of Life Sc 15 • Fatty acyl chain is extended by two carbons, When the chain length reaches 16 carbons, the product Palmitic acid (16:0) The overall process is Dr. Gautam Kr. Dept. of Life Sc 16 • Carbons C-16 and C-15 of the palmitate are derived from the methyl and carboxyl carbon atoms, respectively, of an acetyl-CoA. • Two-carbon units donated by activated malonate • Loss of CO2 at each step Dr. Gautam Kr. Dept. of Life Sc 17 Routes of synthesis of other fatty acids • Fatty acid elongation systems present in the Smooth Endoplasmic Reticulum and in Mitochondria. Acetyl CoA for FA elongation Malonyl CoA for FA elongation • The more active elongation system of the ER extends the 16-carbon chain of palmitoyl-CoA by two carbons, forming stearoyl-CoA (C-18). Dr. Gautam Kr. Dept. of Life Sc 18 Desaturation of fatty acids in vertebrates • The double bond is introduced into the fatty acid chain by an oxidative reaction catalyzed by fatty acyl–CoA desaturase . • In These reactions take place on the lumenal face of the smooth ER • In plants, oleate is produced by a stearoyl-ACP desaturase in the chloroplast stroma that uses reduced ferredoxin as the electron donor. • Two most common monounsaturated fatty acids of animal tissues: Palmitoleate, 16:1(∆9), and Oleate, 18:1(∆9); both of these fatty acids have a single cis double bond between C-9 and C-10. Smooth ER • Thus mammals cannot synthesize linoleate, 18:2(∆9,12), or α-linolenate, 18:3(∆9,12,15). linoleate and linolenate are essentialDr. Gautamfatty Kr.acids Dept. forof Life mammals Sc 19 Biosynthesis of Triacylglycerols (Glyceroneogenesis) • L-glycerol 3-phosphate and Fatty acyl–CoA : two precursors • Glyceroneogenesis is a shortened version of gluconeogenesis, from pyruvate to DHAP • DHAP to glycerol 3- phosphate by cytosolic NAD-linked glycerol 3-phosphate dehydrogenase • Glycerol 3-phosphate is subsequently used in triacylglycerol synthesis Dr. Gautam Kr. Dept. of Life Sc 20 Biosynthesis of Membrane Phospholipids The assembly of phospholipids from simple precursors requires (1) synthesis of the backbone molecule (glycerol or sphingosine); (2) attachment of fatty acid(s) to the backbone through an ester or amide linkage; (3) addition of a hydrophilic head group to the backbone through a phosphodiester linkage; and, in some cases, (4) alteration or exchange of the head group to yield the final phospholipid product. Dr. Gautam Kr. Dept. of Life Sc 21 In eukaryotic cells Phospholipid synthesis occurs primarily on the surfaces of the smooth endoplasmic reticulum and the mitochondrial inner membrane. Dr. Gautam Kr. Dept. of Life Sc 22 Sterols (Cholesterol) • Sterols Have Four Fused Carbon Rings • Sterols are structural lipids present in the membranes of most eukaryotic cells • Much of the cholesterol synthesis in vertebrates takes place in the liver. • Bacteria cannot synthesize sterols; a few bacterial species, however, can incorporate exogenous sterols into their membranes. • The sterols of all eukaryotes are synthesized from simple five carbon isoprene subunits, as are the fat-soluble vitamins, quinones, and dolichols Cholesterol Dr. Gautam Kr. Dept. of Life Sc 23 • The sterols serve as precursors for a variety of products with specific biological activities • Bile acids are polar derivatives of cholesterol that act as detergents in the intestine, emulsifying dietary fats to make them more readily accessible to digestive lipases. Dr. Gautam Kr. Dept. of Life Sc 24 Steroids derived from cholesterol • Testosterone, the male sex hormone, is produced in the testes. Estradiol, one of the female sex hormones, is produced in the ovaries and placenta. • Cortisol and aldosterone are hormones synthesized in the cortex of the adrenal gland • Prednisolone and prednisone are synthetic steroids used as anti inflammatory agents Dr. Gautam Kr. Dept. of Life Sc 25 • Plants Use Steroids, and Eicosanoid like Compounds in Signalling • Brassinolide and the related group of brassinosteroids are potent growth regulators in plants, increasing the rate of stem elongation and influencing the orientation of cellulose microfibrils in the cell wall during growth. • Jasmonate, derived from the fatty acid 18:3 (∆9,12,15) in membrane lipids, is chemically similar to the eicosanoids of animal tissues and also serves as a powerful signal, triggering the plant’s defenses in responseto insect-inflicted damage. Dr. Gautam Kr. Dept. of Life Sc 26 Biosynthesis of Cholesterol • Cholesterol is an essential molecule in many animals, including humans, but is not required in the mammalian diet—all cells can synthesize it from simple precursors. • The structure of this 27-carbon compound suggests a complex biosynthetic pathway, but all of its carbon atoms are provided by a single precursor—acetate • The isoprene units that are the essential intermediates in the pathway from acetate to cholesterol Dr. Gautam Kr. Dept. of Life Sc 27 Dr. Gautam Kr. Dept. of Life Sc 28 Overview of isoprenoid biosynthesis Dr. Gautam Kr. Dept. of Life Sc 29 .