Lecture 36 Lipid Metabolism 1 Fatty Acid Oxidation Ketone Bodies Key Concepts Overview of lipid metabolism Reactions of fatty acid oxidation Energy yield from fatty acid oxidation Formation of ketone bodies
Overview of Lipid Metabolism Carbohydrate metabolism is but one component of energy production and storage. In fact, a much larger percentage of the total energy reserves in animals is lipids in the form of fat deposits. Palmitate (16:0) is a C 16 saturated fatty acid that can be carried through the body as a protein-fatty acid complex: Fat is stored in fat cells (adipocytes). Obesity, especially childhood obesity, can be due to both more fat storage per cell, and to a larger number of adipocytes. In contrast, in normal healthy adults, the onset of old age and reduced metabolic rates leads to weight gain resulting primarily from storing more fat per cell (although adults can also add more fat cells if they become obese).
Fatty acids are stored as triacylglycerols (triglycerides) which are uncharged esters of glycerol: Why does this look familiar? Acetyl CoA is the central intermediate in fat metabolism. Fatty acids consist of a long hydrocarbon chain (mostly saturated carbons) with a terminal carboxylate group. Fatty acid degradation, also called fatty acid oxidation, is similar in many ways to fatty acid synthesis - both the catabolic and anabolic pathways utilize acetyl CoA as the activated carrier of the two carbon product or donor.
Triglycerides obtained from food are cleaved by enzymes called lipases in the intestine to generated freey fatty acids and monoacylglycerols. After passing through the membrane of mucosal cells that line the intestine, enzymes catalyze the formation of triglycerides which are then packaged into large lipid rich particles called chylomicrons. We will look at this in more detail in lecture 37.
Pathway Questions 1. What purpose does fatty acid metabolism serve in animals? Fatty acid oxidation in mitochondria is responsible for providing energy to cells when glucose levels are low. Triacylglycerols stored in adipose tissue of most humans can supply energy to the body for ~3 months during starvation. Fatty acid synthesis reactions in the cytosol of liver and adipose cells convert excess acetyl CoA that builds up in the mitochondrial matrix when glucose levels are high into fatty acids that can be stored or exported as triacylglycerols. Pathway Questions 2. What are the net reactions of fatty acid degradation and synthesis for the C16 fatty acid palmitate? Fatty acid oxidation: Palmitate + 7 NAD + + 7 FAD + 8 CoA + 7 H 2 O + ATP 8 acetyl CoA + 7 NADH + 7 FADH 2 + AMP + 2 Pi + 7 H + Fatty acid synthesis: 8 Acetyl CoA + 7 ATP + 14 NADPH + 14 H + palmitate + 8 CoA + 7 ADP + 7 Pi + 14 NADP + + 6 H 2 O
Pathway Questions 3. What are the key enzymes in fatty acid metabolism? Fatty acyl CoA synthetase enzyme catalyzing the "priming" reaction in fatty acid metabolism which converts free fatty acids in the cytosol into fatty acyl- CoA using the energy available from ATP and PPi hydrolysis. Carnitine acyltransferase I - catalyzes the commitment step in fatty acid oxidation which links fatty acyl-coa molecules to the hydroxyl group of carnitine. The activity of carnitine acyltransferase I is inhibited by malonyl-coa, the product of the acetyl-coa carboxylase reaction, which signals that glucose levels are high and fatty acid synthesis is favored. Acetyl CoA carboxylase - catalyzes the commitment step in fatty acid synthesis using a biotin-mediated reaction mechanism that carboxylates acetyl-coa to form the C 3 compound malonyl-coa. The activity of acetyl CoA carboxylase is regulated by both reversible phosphorylation (the active conformation is dephosphorylated) and allosteric mechanisms (citrate binding stimulates activity, palmitoyl-coa inhibits activity). Fatty acid synthase - this large multi-functional enzyme is responsible for catalyzing a series of reactions that sequentially adds C 2 units to a growing fatty acid chain covalently attached to the enzyme complex. The mechanism involves the linking malonyl-coa to an acyl carrier protein, followed by a decarboxylation and condensation reaction that extends the hydrocarbon chain. Pathway Questions 4. What are examples of fatty acid metabolism in real life? A variety of foods are prominently advertised as "non-fat," even though they can contain a high calorie count coming from carbohydrates. Eating too much of these high calorie non-fat foods (e.g., non-fat bagels) activates the fatty acid synthesis pathway resulting in the conversion of acetyl-coa to fatty acids, which are stored as triacylglycerols. In other words, non-fat does not mean that you won t gain fat!
Olestra is a fat substitute composed of a sucrose molecule with several fatty acids attached. Why is this considered a zero calorie fat molecule? Why does Olestra prevent the absorption of fat soluble vitamins? Orlistat is an anti-obesity drug that has been released onto the market. It binds to and inhibits pancreatic lipase activity in the intestine. Why would this function as an anti-obesity drug? What do you think the treatment regimen consists of, and what might be some of the side effects?
Fatty acids stored in adopose cells are released into the blood in response to hormone signaling by activation of cellular lipases which cleave stored triacylglycerides. The fatty acids provide a rich source of energy for tissues throughout the body when glycogen stores have been depleted, especially during endurance exercise and dieting. What two hormones signal through G protein coupled receptors and stimulate energy mobilization, i.e., glycogen breakdown and fatty acid oxidation (lipolysis)? What peptide hormone signals through a receptor tyrosine kinase and functions to inhibit glycogen breakdown and lipolysis? What is the name of this protein kinase?
Fatty acid carbon atoms are numbered starting at the carboxy terminus. The methyl carbon atom at the distal end is called the omega (ω) carbon. The position of double bonds can be denoted from the carboxy end using a delta ( ) sign followed by a superscript number, e.g. cis- 9 -hexadecenoate, or from the CH 3 terminal end using the omega carbon, e.g., omega-3 (ω-3) fatty acid. Polyunsaturated omega-3 fatty acids are found in fish and can be obtained from nutritional supplements. They may play an important role in cardiovascular function and are recommended by the American Heart Association. There is also some recent evidence that they may improve brain function in some patients with bipolar disorder. Read more about this work by Dr. Andrew Stoll at McLean Hospital in Boston. http://www.psychiatrictimes.com/p991211.html You need to be a little careful eating too much large fish (e.g., tuna, swordfish, shark) because of mercury levels. Farm trout may be a good alternative because they are harvested young and cultured under controlled conditions. But there is still concern about the growth conditions and levels of other environmental pollutants from the water.
Fatty acids have four major physiologic roles: 1. Building blocks of membrane phospholipids. 2. Covalently attached to proteins as targeting signals. 3. Major fuel molecules in most animals. 4. FA derivatives are hormones and intracellular signals. Fatty acids vary in chain length and degree of unsaturation (C=C bonds). A fully saturated fatty acid has no C=C bonds.
Triacylglycerols are very anhydrous and can store more energy with less mass than carbohydrates (glycogen). One gram of anhydrous fat stores SIX TIMES more energy than a gram of hydrated glycogen. A man weighing 155 pounds with a normal amount of stored fat, would have to weigh 240 pounds if that same amount of energy were stored as glycogen. Where is the "extra" 85 pounds coming from in this example? Why might your body weight fluctuate if you suddenly change the relative proportion of carbohydrate, protein and fat in your diet?
Reactions of fatty acid oxidation Stored triacylglycerols are first hydrolyzed by lipases to release glycerol and free fatty acids. Conformational changes in Lipase accommodate fatty acid binding
Glycerol is further metabolized to dihydroxyacetone phosphate (DHAP) following phosphorylation and dehydrogenation: What is the metabolic fate of the dihydroxyacetone phosphate under conditions of low energy charge in the cell? Fatty acids are activated by ATP through an activation process on the outer mitochondrial membrane requiring the enzyme acyl-coa synthetase to form fatty acyl-coa. This reaction is driven by the hydrolysis of inorganic pyrophosphate. What enzyme cleaves PPi and why would this pull the acyl-coa synthetase reaction to the right?
Fatty acyl-coa derivatives are carried across the inner mitochondrial membrane by conjugation to carnitine. The carrier process requires carnitine acyltransferase I (outside) and carnitine acyltransferase II (inside). The two carnitine acyltransferase enzymes (I and II) function on either side of the inner mitochondrial membrane Once inside the mitochondrial matrix, fatty acids are oxidized 2 carbons at a time, releasing acetyl-coa, NADH and FADH 2.
There are four steps in fatty acid oxidation pathway: 1) oxidation, 2) hydration, 3) oxidation, 4) thiolysis. 1 This is why it is called Beta Oxidation. 2 3 There are four steps in fatty acid oxidation pathway: 1) oxidation, 2) hydration, 3) oxidation, 4) thiolysis. 3 4
Taken together with acyl-coa synthetase and carnitine acyltransferase reactions, 6 steps are actually required to release the first acetyl-coa from a fatty acid starting in the cytosol. After this initial set of reactions, each round of FA oxidation releases 2 carbons as acetyl-coa, and produces 1 NADH and 1 FADH 2. It requires 7 rounds of FA oxidation to metabolize palmitate, a C 16 fatty acid. The electron pair from FADH 2 is donated directly to ubiquinone in the electron transport system via the ETF:Q oxidoreductase complex as described in lecture 29.
Fatty Acid Oxidation of C 16 palmitate yields 106 ATP! Let's do the math to calculate the ATP yield Palmitoyl-CoA + 7 FAD + 7 NAD + 7 CoA + 7 H 2 O 8 acetyl-coa + 7 FADH 2 + 7 NADH + 7H + 8 acetyl-coa = 24 NADH + 8 FADH 2 +8ATP[GTP] 31 NADH = 31 x 2.5 ATP = 77.5 ATP 15 FADH 2 = 15 x 1.5 ATP = 22.5 ATP Grand Total: 8 ATP + 77.5 ATP + 22.5 ATP = 108 ATP The net energy yield from palmitate oxidation is only 106 ATP. What reaction required 2 high energy phosphate bonds (the equivalent of 2 ATP)?
Note that ATP synthesis and the oxidation of NADH and FADH 2 yields a large amount of H 2 O (subtracting the investment of H 2 O in beta oxidation). ADP + PO 4 2- ---> ATP + H 2 0 2 NADH + 2 H + + O 2 ---> 2 H 2 0 2 FADH 2 + O 2 ---> 2 H 2 0 In fact, desert animals like the kangaroo rat, derive much of their water from fuel metabolism. The complete oxidation of palmitate generates 145 moles of H 2 O: ~10 mls of H 2 O per gram of palmitate. Formation of ketone bodies Acetyl-CoA derived from fatty acid oxidation enters the Citrate Cycle only if carbohydrate metabolism is properly balanced. When fatty acid oxidation produces more acetyl-coa than can be combined with OAA to form citrate, then the "extra" acetyl-coa is converted to acetoacetyl-coa and ketone bodies, including acetone. Ketogenesis (synthesis of ketone bodies) takes place primarily in the liver. The ketone bodies are exported to other parts of the body, such as the heart muscle, where they are converted back to 2 acetyl-coa units. Why must carbohydrate metabolism be balanced for efficient use of acetyl-coa?
Formation of ketone bodies FAT is the fuel of the carbohydrate furnace - if the furnace is working at capacity, and too much fuel is piling up, then it is converted to ketone bodies and shipped out of the cell! What cyclic pathway functions as the "metabolic furnace" in this analogy? In fasting or diabetes, oxaloacetate (OAA) is consumed to form glucose via the gluconeogenic pathway, therefore, the citric acid cycle is no longer able to function at full capacity.
The acetyl-coa that is accumulating from fatty acid oxidation is used instead to make acetoacetate, acetone, and hydroxybutyrate. While the brain under normal conditions prefers glucose, the heart muscle can metabolize acetoacetate to yield 2 moles of acetyl-coa (the brain can switch over during starvation). Can animals convert fatty acids into glucose? Wait a minute, can't acetyl-coa be used to make OAA, which can then be converted to pyruvate as a substrate for gluconeogenesis?