Yıldırım Beyazıt UniversityMedicalFaculty BiochemistryDepartment Fatty AcidSynthesis Prof. Dr. Fatma Meriç YILMAZ firstname.lastname@example.org
FAT CELL DIET FATTY ACID SYNTHESIS ENERGY
Lipid Digestion • Problems • Lipids are not water soluble • Triglycerides too large to be absorbed • Digestive solution • Triglycerides mix with bile and pancreatic secretions • Emulsification and digestion
Bile • Produced in liver, stored in gallbladder • Alkaline solution composed of: • Bile salts • Cholesterol • Lecithin • Bilirubin • Responsible for fat emulsification • suspentionof small particles in the aqueous environment • Bile salts are amphipathic compounds (containing both hydrophobic andhydrophilic components), synthesized in the liver
Digestion of Lipid • Bile salts emulsify lipids • Pancreatic lipase acts on triglycerides • Triglycerides 2- monoglyceride + 2 fatty acids • Pancreatic colipase • Activated by trypsin • Interacts with triglyceride and pancreatic lipase • Improves activity of pancreatic lipase
Micelle formed by bile salts, triacylglycerols and pancreatic lipase.
Pancreatic Colipase • Secreted from pancreas as procolipase • Activated (cleaved) by trypsin • Anchors lipase to the micelle • One colipase to one lipase(i.e., 1:1 ratio)
Bile Salts Dietary Fat (large TG droplet) Lipase 2-Monoglyceride + 2 FFA Lipid emulsion
Emulsification • Produces small lipid spheres • Greater surface area • Lipases attack TG at 1 and 3 positions Glycerol Fatty Acid1 Glycerol Fatty Acid1 Lipase Fatty Acid2 + Fatty Acid2 2 H20 Fatty Acid3 Fatty Acid3 2 Free Fatty Acids 2-Monoglyceride Triglyceride
Digestion of Lipid • Phospholipase A1 and A2 • Hydrolyzes fatty acids from phospholipids • Cholesterol esterase • Hydrolyzes fatty acids from cholesterol esters
Lipid Absorption • The fatty acids and 2-monoacylglycerols produced by digestion are packaged intomicelles, tiny microdroplets emulsified by bile salts • Mixed micelles move to intestinal mucosal cells (enterocytes) and release contents near cell • The bile salts are re-absorbed further down the gastrointestinal tract (in the ileum), transported to the liver, and finally recycled and secreted back into the digestive tract
Nutrient Absorption - Lipids • Fatty acids, 2-monoglycerides, cholesterol, and cholesterol esters move down concentration gradient (passive diffusion) • Repackaged in intestinal cell for transport to liver • Some is reformed into triglycerides • Chylomicrons • Short- and medium-chain fatty acids (C4 to C12) do not require bile salts for theirabsorption. They are absorbed directly into intestinal epithelial cells. Because theydo not need to be packaged to increase their solubility, they enter the portal bloodand are transported to the liver bound to serum albumin.
In the Enterocyte... • Newly formed triglycerides accumulate as ‘lipid droplets’ at the endoplasmic reticulum • Coated with a protein layer • Stabilizes lipids for transport in lymph and blood(aqueous environment) • Glycerol and short chain fatty acids directly enter mesenteric blood These protein-coated lipid droplets are called chylomicrons
Chylomicrons • particles consisting of phospholipids, triacylglycerols and protein • Apolipoproteins (Apo B48, Apo CII)
Lipid Absorption (Chylomicrons) • Chylomicrons absorbed from enterocytes into lacteals (lymph vessels) • Ultimately enter blood via thoracic duct • Most long chain fatty acids absorbed into lymphatic system • Blood lipids are transported as lipoproteins
Overview of Fatty Acid Uptake • Short- and medium-chain fatty acids • Enter portal blood directly from enterocytes • Bound to albumin in blood • Albumin–FFA complex • Oxidized in liver or elongated and used for triglyceride formation • Long-chain fatty acids • Form chylomicrons • Drain into the lymphatics via the lacteal • Enter bloodstream at the thoracic duct • Slow entry into the blood
Fate of Chylomicrons • Chylomicrons are synthesized in intestinal epithelial cells, secreted into the lymph, pass into the blood, andbecome mature chylomicrons • On capillary walls in adipose tissue and muscle, lipoprotein lipase (LPL) activated by ApoCIIdigests the triacylglycerols (TG) of chylomicrons to fatty acids and glycerol. • Fatty acids (FA) are oxidized in muscle or stored in adipose cellsas triacylglycerols. • The remnants of the chylomicrons are taken up by the liver by receptor-mediated endocytosis. Lysosomal enzymes within thehepatocyte digest the remnants, releasing the products into the cytosol.
Fatty Acid Synthesis • What are some of the differences between fatty acid degradation and synthesis? • location in cell • use of acyl carrier protein vs. coenzyme A • association of synthetic enzymes into complex • use of NADPH as opposed to NAD+ and FAD
Introduction • Comparison of fatty acid synthesis and degradation • Two processes nearly mirror one another
Fatty Acid Synthesis • What is the first committed step in fatty acid synthesis? • formation of malonyl CoA • acetyl CoA carboxylase - biotin
Fatty Acid Synthesis • Intermediates in fatty acid synthesis are linked to an acyl carrier protein • role similar to coenzyme A
Fatty Acid Synthesis • Fatty acid are synthesized and degraded by different pathways. • Synthesis takes place in the cytosol. • Intermediates are attached to the acyl carrier protein (ACP). • In higher organisms, the active sites for the synthesis reactions are all on the same polypeptide. • The activated donor in the synthesis is malonyl–ACP. • Fatty acid reduction uses NADPH + H+. • Elongation stops at C16 (palmitic acid)
Formation of Malonyl CoA • Formation of malonyl–CoA is the committed step in fatty acid synthesis • This reaction is analogous to the pyruvate carboxylase reaction that we saw in gluconeogenesis. • The coenzyme biotin used to activate the carbon dioxide.
Acyl Carrier Protein • The intermediates in fatty acid synthesis are covalently linked to the acyl carrier protein (ACP) • The phosphopantetheine group is like the one found in Coenzyme A. • The ACP is small (77aa), and as whole, is like a large Coenzyme A.
Elongation • In bacteria the enzymes that are involved in elongation are separate proteins; in higher organisms the activities all take place on the same polypeptide. • To start an elongation cycle, Acetyl–CoA and Malonyl–CoA are each transferred to an acyl carrier protein
Elongation • Acyl-malonyl ACP condensing enzyme forms Acetoacetyl-ACP. • This reaction is driven by the decarboxylation. • Essentially the free energy from the ATP that was hydrolyzed to put the CO2 onto the acetyl-CoA when forming the malonyl-CoA is released when the CO2 comes off in the condensation reaction.
Elongation • The next three reactions are similar to the reverse of fatty acid degradation, except • The NADPH is used instead of NADH and FADH2 • The D–enantiomer of Hydroxybutarate is formed instead of the L–enantiomer
Elongation • The elongation cycle is repeated six more times, using malonyl–CoA each time, to produce palmityl–ACP. • A thioesterase then cleaves the palmityl–CoA from the ACP. • The elongation always ends when the fatty acid reaches to 16 carbons
Multifunctional Fatty Acid Synthase-A dimeric enzyme • Domain 1 • Substrate entry and condensation unit • Domain 2 • Reduction unit • Domain 3 • Palmitate release unit
Fatty Acid Synthase Mechanism • Initially, acetyl CoA adds to the synthase. It provides the -methyl groupof palmitate. • Malonyl CoA provides the 2-carbon units that are added to the growing fatty acyl chain. The addition and reduction steps arerepeated until palmitate is produced.
Fatty Acid Synthase Mechanism 1. Transfer of the malonyl group to the phosphopantetheinyl residue. 2. Condensation of the malonyl andfatty acyl groups. 3. Reduction of the -ketoacyl group. 4. Dehydration. 5. Reduction of the double bond.
Stoichiometry of FA synthesis • The stoichiometry of palmitate synthesis: • Synythesis of palmitate from Malonyl–CoA • Synthesis of Malonyl–CoA from Acetyl–CoA • Overall synthesis
Citrate Shuttle • Acetyl–CoA is synthesized in the mitochondrial matrix, whereas fatty acids are synthesized in the cytosol • Acetyl–CoA units are shuttled out of the mitochondrial matrix as citrate:
Citrate Shuttle • The shuttle allows Acetyl-CoA to be shuttled to the cytosol, where fatty acid synthesis can occur. • The shuttle consumes one equivalent of ATP. • The shuttle also substitutes an NADPH for an NADH, which is also needed for synthesis.
Citrate Shuttle • In the cytoplasm, citrate is cleaved to form OAA and Acetyl CoA with the consumption of one ATP • OAA turns to malate and one molecule NADH is used in this step • Pyruvate is synthesized from malate and one NADPH is produced. • Pyruvate enters to the mitochondria and produces OAA
Fatty Acid Synthesis • From where does NADPH needed for synthesis come? • pentose phosphate pathway • Main source • reduction of OAA to malate followed by oxidative decarboxylation of malate to pyruvate
Sources of NADPH • The malate dehydrogenase and NADP+–linked malate enzyme reactions of the citrate shuttle exchange NADH for NADPH
Sources of NADPH • Most of the NADPH for the the synthesis of palmitoil-CoA still comes from the phosphate pentose pathway.
Elongation and Unsaturation • Desaturation of fatty acids involves a process that requires molecular oxygen (O2), NADH, and cytochrome b5. • The process occurs in the endoplasmic reticulum and uses molecular oxygen.
Elongation and Unsaturation • Both the fatty acid and NADHare oxidized. • Human desaturases cannot introduce double bonds between carbon 9 and the methyl end.
Elongation and Unsaturation • Plants are able to introduce double bonds into fatty acids in the region betweenC10 and the methyl end and therefore can synthesize 3 and 6 polyunsaturatedfatty acids. • Fish oils also contain omega 3 and 6 fatty acids, particularly eicosapentaenoic acid (EPA; 3, 20:5, 5, 8, 11, 14, 17) and docosahexaenoic acid (DHA; 3,22:6, 4,7,10,13,16,19). The fish obtain these fatty acids by eating phytoplankton(plants that float in water). • Arachidonic acid is listed in some textbooks as an essential fatty acid. Although it isan omega 6 fatty acid, it is not essential in the diet if linoleic acid is present because arachidonicacid can be synthesized from dietary linoleic acid
Elongation and Unsaturation • Elongation and unsaturation convert palmitoyl–CoA to other fatty acids. • Reactions occur on the cytosolic face of the endoplasmic reticulum. • Malonyl–CoA is the donor in elongation reactions