number 24 Done by Mohammed tarabieh Corrected by Doctor Faisal Al-Khatibe 1 P a g e
*Please look over the previous sheet about fatty acid synthesis **Oxidation(degradation) of fatty acids, occurs in the mitochondria **Reduction(synthesis) of fatty acids, occurs in the cytosol. **These two processes are opposite of each other so each one occurs in a different compartment. _The first step in the synthesis is the carboxylation of acetyl CoA by Acetyl CoA carboxylase to produce malonyl CoA _This is the rate limiting step which is as usual, regulated. Fatty acid de novo synthesis regulations: We are going to be concerned with acetyl CoA carboxylase (ACC) enzyme only, which is the rate limiting enzyme as well. The inactive form of the enzyme (ACC) is a protomer (the structural unit of an oligomeric protein. Composed of a dimer of 2 polypeptide chains). So, for the ACC enzyme to become active it needs to polymerize. Regulation of Acetyl CoA carboxylase is divided into two types:- A. Short-term regulation B. Long-term regulation 2 P a g e
Short-term regulation (activity of the enzyme) Allosterically: - (very rapid within milliseconds) A. Activated by citrate which causes protomers to polymerize. Why citrate? Citrate indicates that the level of energy in the cell is high. And in the case of high energy in the cell citrate accumulates and moves to the cytosol. Citrate also is the precursor of acetyl CoA, it gives Acetyl CoA. B. Inactivated by palmitoyl CoA (the end product of the pathway) which causes depolymerization. Covalent Reversible phosphorylation (longer period of time within seconds/minutes) A. Active when dephosphorylated, occurs by protein phosphatase which is induced by insulin. B. Inactive when phosphorylated, occurs in the presence of hormones such as epinephrine and glucagon (low glucose level in the blood) Phosphorylation of acetyl CoA carboxylase occurs by Adenosine monophosphate-activated protein kinase (AMPK) which is activated: - 1- Allosterically by AMP (low level of energy in the cell so there is no need for fatty acid synthesis. We actually need to degrade Fatty acids) 2- Covalently by phosphorylation via several kinases. One of these AMPK kinases is activated by cyclic AMP dependent protein kinase A (PKA)which is activated by Glucagon and epinephrine. So, protein kinase A (camp dependent) indirectly inactivates Acetyl CoA Carboxylase. In general, adding phosphate to metabolic enzymes favours saving glucose (glucose level is low). 3 P a g e
Long-term regulation (amount of the enzymes) -Diet containing excess calories (high carbohydrates, low fat diets) induces Acetyl CoA carboxylase synthesis. (induction of the enzyme) -Diet containing low calories (low carbohydrates, high fat diets) reduces ACC synthesis. Fatty acid synthase is similarly regulated. The following info is taken from the book, the doctor hasn t mentioned it in any way: - ACC synthesis is upregulated by carbohydrates (specifically glucose) via the transcription factor,carbohydrate response element-binding protein(chrebp)and by insulin via the transcription factor, sterol regulatory element-binding protein-1c (SREBP-1c). Metformin, used in treatment of type 2 diabetes(glucose level is higher than normal ) lowers plasma TAG through activation of AMPK, resulting in inhibition of ACC activity (by phosphorylation) and inhibition of ACC and fatty acid synthase expression (by decreasing SREBP-1c). Metformin lowers blood glucose by increasing AMPKmediated glucose uptake by muscle. Regulation of fatty acid oxidation: Regulation of fatty acid oxidation is related to: - A. Supply of fatty acid B. Hormonal control C. Entry to mitochondria D. Availability of NAD+ Hormone sensitive lipase regulation found in the white adipose tissue 4 P a g e
A. Active when phosphorylated by protein kinase A (camp dependent) which is activated by Glucagon and Epinephrine. B. Inactive when dephosphorylated in the presence of high insulin level Note: when camp pathway is activated we inhibit Acetyl CoA carboxylase (Fatty acid synthesis) and activate hormone sensitive lipase (fatty acid degradation). camp pathway is activated by glucagon and epinephrine which indicates low glucose level. Activity of Hormone sensitive lipase increases the supply of free fatty acid which increases oxidation. Availability of NAD+ Because NAD+ is a substrate needed in each fatty acid cycle where it is converted to NADH. When NADH level is high it means NAD+ level is low, because the total number of NADH and NAD+ is constant. (when one increases the other decreases) When NAD+ level is low fatty acid oxidation is inhibited. Entry to mitochondria (where oxidation occurs) -Carnitine shuttle regulation Malonyl CoA inhibits carnitine acyl transferase I (found on the outer mitochondrial membrane) thus inhibits the entry of long-chain acyl groups into the mitochondrial matrix. When fatty acid synthesis occurs in the cytosol as indicated in the presence of Malonyl CoA, fatty acid degradation is inhibited. 5 P a g e
The idea is that synthesis and degradation should not occur at the same time or else it would just waste energy. Note: short-chain fatty acid oxidation is not inhibited by malonyl CoA as short chain fatty acids move to the mitochondrial matrix without the need for the carnitine shuttle. Acetyl CoA/ CoA ratio As this ratio increases the CoA-requiring thiolase*reaction decreases (degradation of fatty acid decreases) *(different enzymes that require CoA in fatty acid oxidation pathway ex. 3-ketoacyl CoA thiolase) Designing a drug that increases fatty acid oxidation Such a drug would be helpful in combating obesity. Enzymes usually work at their maximum speed, and do not need any external influence (drug) to increase their activity. So, you can inhibit an enzyme and not stimulate an enzyme. Most Drugs usually work by inhibiting the action of an enzyme. What enzyme should we inhibit to increase fatty acid oxidation? Inhibit Acetyl CoA carboxylase -> malonyl CoA decreases and fatty acid oxidation increases. Fatty acid elongation Fatty acid synthase synthesizes fatty acid which are 16 carbons long (palmitate) so how can we get longer fatty acid chains? Further elongation of palmitate occurs in the smooth endoplasmic reticulum. 6 P a g e
Occurs by the addition of 2 carbon units from malonyl CoA to the carboxylate end and requires NADPH as the reducing power. Requires a system of enzymes rather than a multifunctional enzyme. (more than one enzyme is required) Similar sequence of reaction to fatty acid synthase but catalyzed by different enzymes found in the Smooth endoplasmic reticulum Reaction is 4 steps long This elongation process Occurs to fatty acids with 16 carbons. Elongation of fatty acid in the mitochondria Its a different pathway that occurs in the mitochondria (it s a minor pathway) uses the reversal actions of beta oxidation except the first reaction which is Fatty acyl CoA + FAD trans-2-enoyl CoA +FADH2 * this occurs during beta oxidation *Catalyzed by: acyl CoA dehydrogenates So instead of using FADh2 it uses NADPH during elongation 7 P a g e
This pathway occurs to fatty acids shorter than 16 carbons, such fatty acids are found in milk, butter... to store fatty acids shorter than 16 carbons long we need to elongate them to 16 carbons and then we can store them Introduction of double bonds Catalyzed by enzymes (fatty acyl CoA desaturase) introduce cis double bonds.no trans double bonds Occurs in the Smooth endoplasmic reticulum 9 Desaturasereaction requires O2, NADPH, cytochrome b5, and its FAD-linked reductase Fatty acid and NADPH get oxidized as the O2 gets reduced to H2O! (in slides its written NADPH in the book it says NADH)! Fatty acid receives one oxygen atom creating a hydroxyl group at carbon 9 followed by dehydration that introduces double bound not dehydrogenase. The other oxygen atom gets reduced by NADPH to H2O. NADPH is not used to reduce FA We need cytochrome b5 for activation of oxygen. The first double bound is introduced between carbon 9 and 10 producing oleic acid 18:1 (9) and Palmitoleic acid 16:1(9) No double bond can be introduced beyond (after) carbon 9 in humans, no double bound on 12,15 Fatty acids that contain double bonds beyond carbon 9 are essential fatty acids as we can t synthesize them We can introduce double bonds before carbon 9, at carbon 5,6. CH2 must separate between 2 double bonds Formation and Modification of Polyunsaturated FA -Elongation - Desaturation **Additional double bonds can be introduced by: 4 Desaturase 6 Desaturase 5 Desaturase 8 P a g e
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Biosynthesis of Triacylglycerol and phosphoacylglycerol The difference between triacylglycerol and phosphoacyglycerol is that phosophoacyglycerol contains a phosphate group that is esterified to an alcohol on carbon 3. So, since there is similarity between the 2 structures their synthesis will proceed with some common steps. For example, phosphatidic acid (1 glycerol esterified to 2 FA and 1 phosphate) is a common intermediate in both TAG and Phosphoacyglycerol synthesis. Biosynthesis of TAG requires acyl CoA which is the active form of FA and glycerol phosphate not normal glycerol why the active form of FA? why not the free FA? Look at the following reactions The first rxn is a hydrolysis rxn with -ve G. Hydrolysis always proceeds with a large -ve G and is an exergonic rxn because water exists in large amounts The second rxn is opposite of the first and proceeds with a large +ve G; even in the presence of an enzyme. And we can t 10 P a g e
increase FA concentration to a large value to proceed with the rxn The third rxn; energy from breaking high energy bond (thioester bond) in the activated FA makes the rxn proceed with a -ve G. So, we require activated FA to proceed with the rxn. We require 2 ATP to activate FA and synthesize Acyl CoA. This is how the rxn proceeds First rxn; acyl is transferred from Acyl CoA to glycerol 3 phosphate catalyzed by the enzyme acyltranferase to give the compound lysophosphatidic acid (similar to phosphatidic acid but without FA at carbon 2). This is a transfer rxn. Second rxn is similar acyltranferase transfers acyl from acyl CoA to lysophosphatidic acid to produce phosphatidic acid. To transfer the third acyl group and make TAG we need to remove the phosphate group; which is catalyzed by phosphotidate phosphatase by hydrolysis to produce DAG Last step is similar to step 1 and 2 (acyl transfer by acyltranferase) 11 P a g e
How do we get glycerol 3 phosphate? Glycerol kinase is not present in adipose tissue TAG production occurs mainly in two organs: A. Liver: from excess carbohydrates B. Adipose tissue C. Lactating mammary glands *How is TAG synthesized in adipose tissue when they don t contain glycerol kinase? It s produced from dihydroxy acetone phosphate by the action of glycerol phosphate dehydrogenase which oxidizes NADH to NAD+ and reduced Dihydroxy acetone phosphate (ketone) to Glycerol-3-phosphate (alcohol) The significance of glycerol kinase absence in Adipose tissue is simply because; when energy is required TAG is hydrolyzed to make FA and glycerol. FA will move through the circulation to different organs while glycerol will go the liver for conversion to glucose. If glycerol kinase is present in adipose tissue; glycerol that is hydrolyzed for energy will be rapidly phosphorylated and react with FA-CoA to produce TAG again. This synthesis and degradation costs and consumes 7 ATP because we require 3 FA, each one requires 2 ATP, to become in the active form (FA-CoA) =6 ATP and 1 ATP to produce glycerol-3-phosphate. This is prevented by the absence 12 P a g e
of glycerol kinase.so synthesis and degradation are separated. Glucose enters the adipose tissue, and through glycolysis dihydroxy acetone phosphate is produced and used to synthesis Glycerol-3-phosphate, this happens during well fed state when insulin levels in the blood are high. Remember glut-4(insulin dependent) exists in adipose tissue and is induced by insulin. So, insulin is required for glucose to enter adipose tissue. In the case of a diabetic patient insulin is not present and glucose level is high in the blood, glucose will not be able to enter the adipose tissue and the adipose tissue therefore will not be able to synthesis TAG, this leads to a decrease in body weight. A diabetic patient suffers from polyuria(urination) and polydipsia(thirst) which are symptoms of diabetes. How is TAG synthesized in liver? A. By glycerol kinase B. By glycerol phosphate dehydrogenase from dihydroxyacetone phosphate **Ana mn r2ye tshofo the following shows :- 1) The good doctor 2) Shark tank 3) Grand tour season 2 is coming very soon ارطغرل (4 سلطان عبد الحميد (5 13 P a g e