Beta Oxidation and Fatty Acid Synthesis PDF

Topic 8 β oxidation and fatty acid synthesis November 30th, 2021 Topic 8 Fatty acid sources (from food and adipose cell) Beta oxidation Activation 4 Steps Energy yield Odd C number and unsaturated fatty acid oxidation Fatty acid biosynthesis Fatt...

Topic 8 β oxidation and fatty acid synthesis November 30th, 2021 Topic 8 Fatty acid sources (from food and adipose cell) Beta oxidation Activation 4 Steps Energy yield Odd C number and unsaturated fatty acid oxidation Fatty acid biosynthesis Fatty acid synthase enzyme (FAS) Steps Regulation Van Nguyen 1  Lipids: store and generate energy Fats are obtained from Dietary Intake and Adipose Tissue Most of the “fats” in diet and in adipose tissue are triglycerides Hormones (glucagon, epinephrine, ACTH) trigger the release of fatty acids from adipose tissue Acetyl - CoA SYNTHESIS HYDROLYSIS HYDROLYSIS OXIDATION ATP Acetyl - CoA Van Nguyen 2 Fatty acids store and generate high energy The energy yield per gram of fatty acid oxidized is greater than that per gram of carbohydrate oxidized Carbon chains: mostly highly reduced (-CH2-) so its oxidation yields the most energy possible Fatty acids can pack more closely in storage tissues Van Nguyen 3 From dietary intake Hydrolyzed by enzymes Lipases catalyze hydrolysis of triacylglycerols Phospholipases catalyze hydrolysis of phosphoacylglycerols. May have multiple sites of action as shown below Van Nguyen 4  From dietary intake Hydrolysis of triacylglycerols by pancreatic and intestinal lipases. Pancreatic lipases cleave fatty acids at the C-1 and C-3 positions. Resulting monoacylglycerols with fatty acids at C-2 are hydrolyzed by intestinal lipases. Van Nguyen 5 The Liberation of Fatty Acids in adipose cell Fatty acids degradation Knoop showed that fatty acids must be degraded by removal of 2-C units Albert Lehninger showed that this occurred in the mitochondria F. Lynen and E. Reichart showed that the 2-C unit released is acetyl-CoA, not free acetate The process begins with oxidation of the carbon that is "!" to the carboxyl carbon, so the process is called "!-oxidation" Van Nguyen 7  Fatty Acid Activation Fatty acid oxidation begins with activation A thioester bond is formed between the acyl group of the FA and the thiol of CoA-SH Fatty Acid Activation - Mechanism Van Nguyen 9  Fatty Acid Activation – Energy Formation of a CoA ester is expensive energetically Free energy change just barely breaks even with ATP hydrolysis But subsequent hydrolysis of PPi drives the reaction strongly forward Energy spent = (1 ATP + 1 PPi) ~ 2 ATP Van Nguyen 10  The Role of Carnitine in Acyl-CoA Transfer The acyl-CoA crosses the outer mitochondrial membrane (but not the inner membrane) Outer membrane The acyl group is then transferred to carnitine, carried across the inner mitochondrial membrane Carnitine Palmitoyltransferase Inner (CPT-1) has specificity for acyl membrane groups between 14 and 18 carbons long In the matrix, acyl-carnitine exchanges with mitochondrial CoA-SH to reform Acyl-CoA Van Nguyen 11 b-Oxidation b-Oxidation: a series of reactions that cleaves carbon atoms two at a time from the carboxyl end of a fatty acid The complete cycle of one b-oxidation requires four enzymes Reaction 1: Oxidation of the a,b carbon-carbon single bond to a carbon-carbon double bond Reaction 2: Hydration of the carbon-carbon double bond Reaction 3: Oxidation of the b-hydroxyl group to a carbonyl group Reaction 4: Cleavage of the carbon chain by a reverse Claisen reaction Van Nguyen 12 b-Oxidation Van Nguyen 13 Step 1: acyl-CoA dehydrogenase reaction Removal of a proton from the !-carbon is followed by hydride transfer from the "-carbon to FAD. Van Nguyen 14 Step 2: enoyl-CoA hydratase or crotonase reaction Stereospecific: The enzyme converts trans-enoyl-CoA derivatives to L-"-hydroxyacyl-CoA. Van Nguyen 15 Step 3: L-hydroxyacyl-CoA dehydrogenase reaction Van Nguyen 16 Step 4: !-ketothiolase reaction Attack by an enzyme’s Cys thiolate group at the "-carbonyl carbon produces a tetrahedral intermediate, which decomposes with departure of acetyl-CoA, leaving an enzyme thioester intermediate. Attack again by the thiol group of a second CoA yields a new acyl-CoA. Van Nguyen 17 Summary Fatty acids are activated and transported to the mitochondrial matrix for further catabolism The breakdown of fatty acids takes place in the mitochondrial matrix and proceeds by successive removal of two-carbon units as acetyl-CoA Each Cleavage of a two-carbon moiety requires a four-step reaction sequences called b-oxidation Van Nguyen 18 Ketone Bodies Formed when acetyl-CoA exceeds oxaloacetate amount available (to react in the citric acid cycle) Intake high in lipids and low in carbohydrates Diabetes not suitably controlled Starvation Ketone bodies are: acetone, b-hydroxybutyrate &acetoacetate Formed principally in liver mitochondria Can be used as a fuel in most tissues and organs Van Nguyen 19 Number of cycles and number of 2-Carbon units 8 cycles of b-Oxidation of Steric acid (18 C) produces 9 (2-Carbon) Units Van Nguyen 20  Energy Yield from Fatty acid Oxidation Molecules Activation Beta oxidation TCA, (e transport, Total Convert to step step ATP synthesis) ATP ATP -2 -2 -2 NADH +8 +27 +35 +87.5 FADH2 +8 +9 +17 +25.5 GTP +9 +9 +9 CoA-SH -1 -8 +9 0 0 1 NADH = 2.5 ATP 1 FADH2 = 1.5 ATP 1 GTP = 1 ATP 1 Molecule of stearic (18 C) acid: +120 ATP Step Spend (-) Produce (+) Activation - 2 ATP, - 1 CoA-SH Beta oxidation - (n/2-1) CoA-SH +(n/2-1) NADH, +(n/2-1) FADH2, TCA + 3n/2 NADH, + n/2 FADH2, + n/2 GTP, + n/2 CoA-SH Van Nguyen 21 Energy Yield from Fatty acid Oxidation Molecules Activation Beta oxidation TCA, (e transport, Total Convert to step step ATP synthesis) ATP ATP NADH FADH2 GTP CoA-SH 1 Molecule of palmitic (16 C) acid: ? Van Nguyen 22 Energy Yield from Fatty acid Oxidation Molecules Activation Beta oxidation TCA, (e transport, Total Convert to step step ATP synthesis) ATP ATP -2 -2 -2 NADH +7 +24 +31 +77.5 FADH2 +7 +8 +15 +22.5 GTP +8 +8 +1 CoA-SH -1 -7 +8 0 0 1 Molecule of palmitic (16 C) acid: +106 ATP Van Nguyen 23  Energy Yield from Fatty acid Oxidation ATP Free energy ~ - 30.5 kJ/mol 120 (mol) ATP ~ 3660 kJ ~ 875 kcal This high energy release (and their low density) makes fatty acids the fuel of choice for migratory birds and many other animals Fun estimation: A bird weighs 200 g with 10% fat needs ~ 7.5 kcal/day. Assume all fat is equivalent to stearic acid. Estimate the number of days the bird can survive using only its fat as the source of energy. (Ignore all other conditions). Van Nguyen 24  Oxidation of Odd-Numbered FA The last β-oxidation cycle of a fatty acid with an odd number of carbons (not frequently encountered) gives propionyl-CoA  Oxidation of an Unsaturated FA A cis-trans isomerization is needed to convert unsaturated FA to acetyl-CoA This enzyme is known as an isomerase Oxidation of unsaturated FA does not generate as much ATP relative to saturated FA with the same # of carbons Van Nguyen 26 Summary FA with odd number of carbons produce propionyl-CoA in the last step of the oxidation Propionyl-CoA can be converted to succinyl-CoA, which plays a role in the citric acid cycle The oxidation of unsaturated FA requires enzymes that catalyze isomerization around the double bonds so that oxidation can proceed Van Nguyen 27 Fatty Acid Biosynthesis Acyl-CoA is transferred into the mitochondria by Carnitine carrier. Acyl-CoA (nC) will be oxidized/broken down to Acetyl-CoA (2C), which is then transported out to the cytosol. Fatty acid biosynthetic reactions occur in the cytosol Fatty Acid Biosynthesis Steps 1. Acetyl-CoA (2C) + HCO3- à Malonyl-CoA (3C) 2. Acetyl-CoA (2C) + Malonyl-CoA (3C) à à… Acetoacetyl-ACP (4C) + CO2. 3. Reduction of beta-carbonyl group 4. Dehydration to form α, β double bond C=C 5. Reduction of double bond to form saturated C chain Acetoacetyl-ACP (4C) à Butyryl-ACP (4C) 6. Butyryl-ACP (4C) + Malonyl-CoA (3C) à … [repeat step 3 to 5] à Hexanoyl-ACP (6C) + CO2. … n. [repeat step 3 to 5] à Acyl-ACP (nC) Step 1: Acetyl-CoA Carboxylase (ACC) Carboxylation of acetyl-CoA occurs in the cytosol Catalyzed by acetyl-CoA carboxylase Biotin is an important cofactor, the carrier of the carboxyl group Malonyl-CoA is key intermediate that is produced ACC enzyme Step 1: Acetyl-CoA Carboxylase Mechanism Fatty Acid Synthase (FAS) Van Nguyen 32 Step 2: β-ketoacyl-ACP synthase Acetyl activation S-CoA Van Nguyen 33 Step 2: β-ketoacyl-ACP synthase - Malonyl activation S-CoA Van Nguyen 34 Step 2: β-ketoacyl-ACP synthase Condensation Decarboxylation drives the β-ketoacyl-ACP synthase and results in the addition of two-carbon units to the growing chain. Van Nguyen 35  Step 3-5: 3. Reduction (KR enzyme) 4. Dehydration (HD enzyme) 5. Reduction (ER enzyme) Van Nguyen 36 Step 6 to n: Repeat adding 2 C l o cation Trans Newly synthesized fatty acids exist primarily as acyl-CoA esters à Concentrations of free fatty acids areVan extremely Nguyen low in most cells 37  Biosynthesis of Van Nguyen Palmitate from Acetyl- and Malonyl-CoA 38 Biosynthesis of Palmitate from Acetyl- and Malonyl-CoA Van Nguyen 39 Summary Acetyl-CoA is transported to the cytosol and converted to malonyl-CoA The biosynthesis of FA proceeds by the addition of 2-carbon units to the hydrocarbon chain. The process is catalyzed by the fatty-acid synthase complex Comparison of FA Degradation and Biosynthesis Sites of Fatty Acid Metabolism in an Animal Cell Van Nguyen 41 Palmitate synthesis from Acetyl-CoA Van Nguyen 42 Regulation Van Nguyen 43 Regulation Van Nguyen 44 References Viel & Gaudet, Principle of Biochemistry, 2015 Garrett & Grisham, Biochemistry, 2017 Campbell & Farrell, Biochemistry, 2016 Lehninger, Principle of Biochemistry, 2013 Van Nguyen 45