Clinical biochemistry / second stage Dr.Khawla A. Shemran



Fatty acid Synthesis Mechanism:Acetyl-CoA Carboxylase, which converts acetyl-CoA to malonyl-CoA, is the committed step of the fatty acid synthesis pathway. The acetyl-CoA and malonyl-CoA are transferred to Acyl carrier protein (ACP) by the action of acetyl-CoA transacylase and malonyl-CoA transacylase, respectively. The attachment of these carbon atoms to ACP allows them to enter the fatty acid synthesis cycle. The synthesis of fatty acids from acetyl-CoA and malonyl-CoA is carried out by fatty acid synthase (FAS). Note that this reaction is an energy requiring process (1 ATP per Malonyl-CoA formed).This multifunctional enzyme catalyzes the seven different reactions whereby two carbon units from malonyl-CoA are linked together, ultimately to form palmitoyl-CoA. In some systems, the activities are present on separate enzyme units.The overall synthesis of palmitate from acetyl-CoA requires 14 NADPHs, and 7 ATPs.Initiation Stage:- The elongation phase of fatty acid synthesis starts with the formation of acetyl ACP and malonyl ACP. Acetyl transacylase and malonyl transacylase catalyze these reactions.Acetyl CoA + ACP acetyl ACP + CoA Malonyl CoA + ACP malonyl ACP + CoAStep 1: loading of acetyl-CoA onto fatty acid synthase Step 2: loading of malonyl- CoA onto fatty acid synthase Steps of fatty acid synthesis starting with Acetyl-CoA and Malonyl-CoA are shown in in the given Figure.The reactions are as follows:Transfer of the malonyl group of malonyl-CoA to Acyl carrier protein (ACP) (Reaction (1) - catalyzed by malonyl-CoA-ACP transacylase). Transfer of the acetyl group of Acetyl-CoA to ACP (Reaction (2) of -catalyzed by acetyl- coenzyme A-acyl-carrier-protein transacylase (acetyl-CoA-ACP transacylase)).Addition of an acetyl group from malonyl-ACP between the thioester bond of the acetyl-ACP molecule in reaction (1) (Reaction (3) of – catalyzed by Beta-keto-ACP synthase – also called condensing enzyme).Reduction of the Beta-keto group to a Beta-hydroxyl group with NADPH (Reaction (4) of – catalyzed by Beta-keto-ACP reductase).Dehydration between the alpha and Beta carbons (Reaction (5) of – catalyzed by Beta-hydroxyacyl-ACP dehydratase).Reduction of the trans double bond by NADPH (Reaction (6) of – catalyzed by enoyl-ACP reductase).Repetition of steps (2-6) six more times. The acetyl group of reaction 1 is replaced by the growing acyl-ACP molecule. (That is, new acetyl groups are added at the ACP end of the molecule).The product of this series of reactions, palmitoyl-ACP can be cleaved to palmitate and ACP by the enzyme palmitoyl thioesterase.Overall reaction of palmitate synthesis from acetyl CoA and malonyl CoA*Acetyl CoA + 7 Malonyl CoA + 14 NADPH + 14 H+ *Palmitate + 7 CO2 + 14 NADP+ + 8 HS-CoA + 6 H2O Elongation of fatty acid chainsAlthough palmitate, is the primary end-product of fatty acid synthase activity, it can be further elongated by the addition of two-carbon units in the endoplasmic reticulum (ER) and the mitochondria. These organelles use separate enzymic processes. elongation of very long chain fatty acids (ELOVL), 3-ketoacyl-CoA reductase (HSD17B12), 3-hydroxyacyl dehydratase (HACD) and trans-2,3,-enoyl-CoA reductase (TECR).The first step in this reaction is catalyzed by the enzyme referred to as “elongation of very long-chain fatty acids” (ELOVL). Seven elongases have been identified in mammals and are designated ELOVL1-7. Desaturations of fatty acid chainsEnzymes present in the ER are responsible for desaturating fatty acids (that is, adding cis double bonds). Termed mixed-function oxidases, the desaturation reactions require NADPH a variety of polyunsaturated fatty acids (PUFA) can be made through additional desaturation combined with elongation.**[Note: Humans lack the ability to introduce double bonds at carbon 9 therefore, must have the polyunsaturated linoleic and linolenic acids provided in the diet .Storage of fatty acids as components of triacylglycerolsMono-, di-, and triacylglycerols consist of one, two, or threemolecules of fatty acid esterified to a molecule of glycerol. Fattyacids are esterified through their carboxyl groups, resulting in a lossof negative charge and formation of "neutral fat." The three fatty acids esterified to a glycerol are usually not of the same type. A fatty acid must be converted to its activated form (attached to Coenzyme A (CoA) before it can participate in TAG synthesis. This reaction is catalyzed by a family of fatty acyl CoA synthetases (thiokinases).Synthesis of a molecule of TAG from glycerol phosphate and fatty acyl CoA involves four reactions; include the sequential addition of two fatty acids from fatty acyl CoA, the removal of phosphate, and the addition of the third fatty acid.Pathways for triacylglyceride synthesis.Triacylglycerides (triglycerides) are synthesized by virtually all cells. The major tissues for TAG synthesis are the small intestine, the liver, and adipocytes. Except for the intestine and adipocytes, TAG synthesis begins with glycerol. In the liver, glycerol is first phosphorylated by glycerol kinase and then activated fatty acids (fatty acyl-CoA's) serve as substrates for fatty acid addition generating phosphatidic acid. The phosphate group is then removed and the last fatty acid is added. In the small intestine, dietary TAGs are hydrolyzed to free fatty acids and monoacylglycerides (MAGs) prior to uptake by the enterocytes. The enterocyte MAGs serve as substrates for acylation in a two-step process yielding a TAG. Within adipose tissue there is no expression of glycerol kinase so the building block for TAG in this tissue is the glycolytic intermediate, dihydroxyacetone phosphate, DHAP. The DHAP is reduced to glycerol-3-phosphate by cytosolic glycerol-3-phosphate dehydrogenase and the remaining reaction of TAG synthesis are the same as for all other tissues. The glycerol backbone of TAGs is activated by phosphorylation at the C-3 (sn-3) position by glycerol kinase. The acylation of glycerol-3-phosphate is the major pathway for triglyceride synthesis in most human cells. Once formed, glycerol-3-phosphate is acylated at the sn-1 position by one of a family of glycerol-3-phosphate acyltransferase (GPAT) enzymes that are expressed in either the mitochondria or in the endoplasmic reticulum, ER. The products of the acylation reactions are lysophosphatidic acids, LPAs. Lysophosphatidic acids are themselves potent bioactive lipidsIntestinal monoacylglycerides (MAG), derived from the hydrolysis of dietary fats, can also serve as substrates for the synthesis of triglycerides (TAGs) which can then be incorporated into chylomicrons. The intestinal synthesis of a TAG from a MAG first involves MAG acylation carried out by one of a family of monoacylglycerol O-acyltransferases (MOGAT). The phosphate of phosphatidic acid is removed, by phosphatidic acid phosphatase (PAP1), to yield 1,2-diacylglycerols (DAG). The fatty acids incorporated into TAGs are all activated to acyl-CoAs through the action of various acyl-CoA synthetasesAdipocytes lack glycerol kinase and, therefore, triglyceride synthesis requires the utilization of DHAP for the backbone in these cells. Within the adipocyte cytosol, DHAP is converted to glycerol-3-phosphate via the action of the cytoplasmic version of glycerol-3-phosphate dehydrogenase .Following formation of a lysophosphatidic acid, various acylglycerol-3-phosphate acyltransferases (AGPAT) add another CoA-activated fatty acid to the sn-2 position generating 1,2-diacylglycerol phosphates which are commonly identified as phosphatidic acids (PA). All of the AGPAT enzymes are integral membrane proteins with the AGPAT1 and AGPAT2 enzymes specifically localized to the ER. DHAP can also serve as a backbone precursor for TAG synthesis in tissues other than adipose, but does so to a much lesser extent than glycerol. In both liver (the primary site of TAG synthesis) and adipose tissue, glycerol phosphate can be produced from glucose, using first the reactions of the glycolytic pathway to produce dihydroxyacetone phosphate (DHAP). Next, DHAP is reduced by glycerol phosphate dehydrogenase to glycerol phosphate. A second pathway found in the liver, but NOT in adipose tissue, uses glycerol kinase to convert free glycerol to glycerol phosphate. And adipocytes lack glycerol kinase, therefore, dihydroxyacetone phosphate (DHAP), produced during glycolysis, is the precursor for TAG synthesis in adipose tissue. This means that adipocytes must have glucose to oxidize in order to store fatty acids in the form of TAGs. ................
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