Class Notes (835,673)
Canada (509,326)
Biology (Sci) (2,472)
BIOL 201 (261)
Lecture 1

BIOL 201 Lecture 1: Cell Biology and Metabolism Part I

81 Pages
Unlock Document

Biology (Sci)
BIOL 201
Gary Brouhard

Thursday, January 7, 2016 Week 1 Lecture 1. Energy in Biological Systems I. - Lodish et al., pp. 43‐45; 49‐51; Berg et al., pp. 427‐429. - E.Coli is a very simple organism; it grows and divides quickly. It requires requires a carbon source (Glucose), NH , 4O and 4alts. It must convert all those into amino acids, nucleotides, lipids, sugars, vitamins (coenzymes) and macromolecules (DNA, RNA, proteins, polysaccharides). The synthesis and breakdown occurs through interconnected pathways involving many different reactions called the metabolic pathway. - Cellular reactions are governed by rules that govern all chemical reactions. Each reaction is catalyzed by specific enzymes binding specific ligands. - Organisms devote much of their genome to specifying metabolic proteins (see table on the right). A lot of genes have metabolic function. - Basic principles of a cell’s metabolism? 1. Fuels are degraded and large molecules are constructed step by step in a series of linked reactions called metabolic pathways. 2. An energy currency common to all life forms, adenosine triphosphate (ATP), links energy-releasing pathways with energy-requiring pathways. 3. The oxidation of carbon fuels powers the formation of ATP. 4. Although there are many metabolic pathways, a limited number of types of reactions and particular intermediates are common to many pathways. 5. Metabolic pathways are highly regulated. 1 Thursday, January 7, 2016 Lecture 2. Energy in Biological Systems II. - Lodish et al., pp. 51 – 52, 78‐81; Berg et al. pp. 429‐430. - An enzyme’s active site consists of two functionally important regions: the substrate binding site, which recognizes and binds the substrate or substrates, and the catalytic site, which carries out the chemical reaction once the substrate has bound. - Reactions that release energy are exergonic (release energy) and catabolic (breaking down). On the other hand, endergonic reactions are anabolic (building up). - What is energy? Energy is the ability to do work from a scientific standpoint. - Energy can be kinetic (muscle contraction) or potential ( chemical bonds , concentration gradients , charge separation across membranes) and energy can be converted from one form into another. - All chemical reactions are reversible. At equilibrium, the forward and reverse rates are the same. The equilibrium constant is characteristic of a reaction. 2 Thursday, January 7, 2016 - We don’t really define the Gibb’s free energy (G), we care about the change in free energy. According to the picture on the right, ΔG dictates the side of the reaction: • if ΔG<0, the forward reaction is favoured • if ΔG>0, the reverse reaction is favoured • if ΔG=0, the system is at equilibrium - An important thermodynamic fact is that the overall free-energy change for a chemically coupled series of reactions is equal to the sum of the free-energy changes of the individual steps. In other words, standard free energy values are additive. Therefore It is possible to form higher energy compounds like glutamine from lower energy compounds like glutamate and ammonia by coupling their synthesis to energy yielding (exergonic) reactions like the hydrolysis of ATP. 3 Thursday, January 7, 2016 0 - The standard free energy change is ΔG . In the equation on the bottom, 2.303 is the factor to convert the “ln" to “log”. - The table below shows the relationship between standard free energy and the equilibrium constant. Based on the previous picture and this table, we observe a positive change in free energy (ΔG>0) if [C][D]/[A][B] at the moment is greater than the equilibrium constant meaning that there are more products than reactants. Therefore, this explains why the reverse reaction is favoured when ΔG>0. 4 Monday, January 11, 2016 Week 2 Lecture 3. Energy in Biological Systems. - Berg et. al. 430‐437. - ATP is the currency of energy within the cell. It is called a high-energy phosphate (high ∆G meaning that it has lots of free energy to release since the products are a lot more stable than the starting material) compound for three main reasons: • ADP and the inorganic phosphate have more resonance stabilization compared to ATP which lowers the free energy. • At pH 7, the triphosphate unit of ATP carries about four negative charges. These charges repel one another because they are in close proximity. The repulsion between them is reduced when ATP is hydrolyzed; therefore, electrostatic repulsion lowers the free energy. • More water can bind more effectively to ADP and P than can bind to the phosphoanhydride part of ATP, stabilizing the ADP and P by hydration. i 1 Monday, January 11, 2016 - For a reaction to occur, reacting species must pass through a higher energy state, the “transition state”. Catalysts act to accelerate the rates of reactions by reducing the ΔG between the reactants and the transition state. Enzymes are natural catalysts: • Enzymes are specific catalysts for biological reactions (key&lock model). An enzyme has a substrate binding site and a catalytic site; together they form the active site. • Enzymes act by binding to a reactant or reactants (termed substrate[s]) in a way that reduces the energy required to reach the transition state. • Enzymes affect only the rates of the reaction – other properties (e.g. K , ΔG) remain the same. eq Enzymes return to their initial state once the • reaction is complete. • E + S ES —> E + P 2 Monday, January 11, 2016 - It is important to notice that he smaller the K m the higher the affinity the enzyme has for the substrate. - Photosynthetic organisms capture light energy and use it to: • transfer the H atoms from water to acceptor molecules • form molecular oxygen • synthesize ATP from ADP and phosphate • transfer H atoms from acceptor to carbons derived from CO to for2 glucose (carbon fixation) • Net: 6 H 2 + 6 CO —>2C H O +66 12 6 2 - Other organisms feed on photosynthetic organisms (directly, indirectly): • Sugars, derived compounds formed through photosynthesis acquired and oxidized to water, carbon dioxide = respiration • Respiration coupled to the formation of ATP from ADP and phosphate (oxidative phosphorylation) • Ultimate electron acceptor is O ,2oxidation product is CO 2 - The ultimate source of all biological energy is sunlight. 3 Monday, January 11, 2016 4 Monday, January 11, 2016 - In metabolism, there are lots of cycles: • NADH and FADH2 reoxidized to NAD and FAD by the respiratory chain • Electrons transferred to O to2form water (end of the electron transport chain) • Lots of free energy released: captured as proton gradient across the mitochondrial inner membrane • The free energy of the proton gradient used to drive the synthesis of ATP from ADP and phosphate • All this together is oxidative phosphorylation - NAD /NADH and FAD/FADH high turnover rates: continually oxidized and reduced during foodstuff oxidation. They are electron carriers. • Metabolism is dynamic • Foodstuffs constantly broken down to carbon dioxide and water + + • NAD gets reduced to NADH then oxidized back to NAD with P + ADP —> ATP.i ATP hydrolysis back to ADP and P used io drive cellular processes. 5 Monday, January 11, 2016 Lecture 4. Glycolysis. - Berg et al., pp. 453‐469. - Glycolysis is the sequence of reactions that metabolizes one molecule of glucose to two molecules of pyruvate with the concomitant net production of two molecules of ATP. This is an anaerobic process taking place in the cytosol. - Glycolysis is an example of fermentation (metabolic process with organic compound as end product). In muscles, the product of fermentation is lactic acid, in yeast, it’s ethanol. Glycolysis is an ancient metabolic pathway and is found in most organisms. - In the first stage of glycolysis, we invest 2 ATP but 4 ATP (2x2 ATP) is gained in the second stage so there’s a net gain of 2 ATP. - Kinases are enzymes that transfer γ phosphoryl group of ATP to acceptor. 6 Monday, January 11, 2016 7 Monday, January 11, 2016 1. Hexokinase uses an ATP to phosphorylate glucose since glucose 6-phosphate cannot pass through the membrane because it is not a substrate for the glucose transporters, and the addition of the phosphoryl group acts to destabilize glucose, thus facilitating its further metabolism. 2. The isomerization of glucose 6-phosphate to fructose 6-phosphate is a conversion of an aldose into a ketose. The reaction catalyzed by phosphoglucose isomerase takes several steps because both glucose 6-phosphate and fructose 6-phosphate are present primarily in the cyclic forms. The enzyme must first open the six- membered ring of glucose 6-phosphate, catalyze the isomerization, and then promote the formation of the five-membered ring of fructose 6-phosphate. 3. Fructose 6-phosphate is phosphorylated at the expense of ATP to fructose 1,6- bisphosphate. This reaction is catalyzed by phosphofructokinase (PFK) and is very important since it is a rate regulating step. 4. The newly formed fructose 1,6-bisphosphate is cleaved into glyceraldehyde 3- phosphate (GAP) and dihydroxyacetone phosphate (DHAP) by the enzyme aldolase. Glyceraldehyde 3-phosphate is on the direct pathway of glycolysis, whereas dihydroxyacetone phosphate is not. These compounds are isomers that can be readily interconverted: dihydroxyacetone phosphate is a ketose, whereas glyceraldehyde 3-phosphate is an aldose. 5. The isomerization of these three-carbon phosphorylated sugars is catalyzed by triose phosphate isomerase. At equilibrium, 96% of the triose phosphate is dihydroxyacetone phosphate. However, the reaction proceeds readily from dihydroxyacetone phosphate to glyceraldehyde 3-phosphate because the subsequent reactions of glycolysis remove this product. The preceding steps in glycolysis have transformed one molecule of glucose into 6. two molecules of glyceraldehyde 3-phosphate, but no energy has yet been extracted. On the contrary, thus far, two molecules of ATP have been invested. We come now to the second stage of glycolysis, a series of steps that harvest some of the energy contained in glyceraldehyde 3-phosphate as ATP. The initial reaction in this sequence is the conversion of glyceraldehyde 3-phosphate into 1,3- bisphosphoglycerate (1,3-BPG), a reaction catalyzed by glyceraldehyde 3- phosphate dehydrogenase. This reaction can be viewed as the sum of two + processes: the oxidation of the aldehyde to a carboxylic acid by NAD andthe joining of the carboxylic acid and orthophosphate to form the acyl-phosphate product. 8 Monday, January 11, 2016 7. 1,3-Bisphosphoglycerate is an energy-rich molecule with a greater phosphoryl- transfer potential than that of ATP. Thus, 1,3-BPG can be used to power the synthesis of ATP from ADP. Phosphoglycerate kinase catalyzes the transfer of the phosphoryl group from the acyl phosphate of 1,3-bisphosphoglycerate to ADP. ATP and 3-phosphoglycerate are the products. The formation of ATP in this manner is referred to as substrate-level phosphorylation because the phosphate donor, 1,3- BPG, is a substrate with high phosphoryl-transfer potential. We will contrast this manner of ATP formation with the formation of ATP from ionic gradients. 8. The position of the phosphoryl group shifts in the conversion of 3-phosphoglycerate into 2-phosphoglycerate, a reaction catalyzed by phosphoglycerate mutase. In general, a mutase is an enzyme that catalyzes the intramolecular shift of a chemical group, such as a phosphoryl group. 9. The dehydration of 2-phosphoglycerate introduces a double bond, creating an enol. Enolase catalyzes this formation of the enol phosphate phosphoenolpyruvate (PEP). This dehydration markedly elevates the transfer potential of the phosphoryl group. An enol phosphate has a high phosphoryl-transfer potential, whereas the phosphate ester of an ordinary alcohol, such as 2-phosphoglycerate, has a low one. 0 The ∆G of the hydrolysis of a phosphate ester of an ordinary alcohol is -13 kJ/mol, whereas that of phosphoenolpyruvate is -62 kJ/mol. 10. The phosphoryl group traps the molecule in its unstable enol form. When the phosphoryl group has been donated to ATP, the enol undergoes a conversion into the more stable ketone —> pyruvate. The virtually irreversible transfer of a phosphoryl group from phosphoenolpyruvate to ADP is catalyzed by pyruvate kinase. 9 Monday, January 11, 2016 -The conversion of glucose into two molecules of pyruvate has resulted in the net synthesis of ATP. However, an energy-converting pathway that stops at pyruvate will not proceed for long, because redox balance has not been maintained. NAD must be regenerated for glycolysis to proceed. Thus, the final process in the pathway is the regeneration of NAD through the metabolism of pyruvate. - When the lactic acid builds up, the pH in the cell decreases and muscles get tired. 10 Monday, January 11, 2016 Lecture 5. Regulation of glycolysis and gluconeogenesis. - Berg et al., pp. 472‐88. - During glycolysis, enzymes that catalyze irreversible steps are allosterically regulated: • Phosphofructokinase ( PFK) is inhibited by ATP, citrate, H , and activated by AMP , fructose-2,6-bisphosphate (F-2,6-BP). • Hexokinase is inhibited by its end product —> glucose-6-phosphate. • Pyruvate kinase is is inhibited by ATP and activated by fructose-1,6-bisphosphate. - PFK is the key regulator of this metabolic pathway. This enzyme is allosterically regulated and it has 4 identical subunits and each subunit has a catalytic site which can bind ATP and fructose-6-phosphate. • PFK has 2 ATP binding sites: 1 catalytic, 1 regulatory. The graph on the right shows the enzyme’s activity depending on the concentrations of ATP and fructose-6-phosphate. • ATP bound at the regulatory site lowers affinity for fructose-6-phosphate. AMP reverse inhibition by ATP; AMP acts as a positive regulator. There’s an enzyme called adenylate kinase (picture on the right) and it creates AMP; when AMP concentration is high, the cells know that there is a huge lack of ATP and will turn phosphofructokinase back on. • Citrate (citric acid) enhances ATP inhibition (liver only). TCA occurs in the mitochondria, if it ends up in the cytosol, it means that there is too much ATP. • PFK is also inhibited by low pH (high proton concentration) and prevents excessive accumulation of lactic acid. • Fructose-2,6,-bisphosphate is a key regulator of PFK (liver only). When glucose levels are high in the blood, fructose-6-phosphate levels are also high, which stimulates a second enzyme called phosphofructokinase-2 (PFK2). PFK2 creates fructose-2,6,-bisphosphate and this molecule decreases the inhibitory effect of ATP, in other words, it increases PFK’s affinity for fructose-6-phosphate and enhances glycolysis. 11 Monday, January 11, 2016 - Additional regulation: hexokinase and pyruvate kinase: • At rest, glucose-6-phosphate will accumulate and shut off hexokinase. Glucose-6- phosphate can also be converted into glycogen for storage. PFK is also shut off by high levels of ATP which also shuts off pyruvate kinase. • During exercise, there is less ATP present and more AMP. Hexokinase will be active since AMP activates PFK which stimulates hexokinase to produce more glucose-6-phosphates. The high level of fructose-1,6-bisphophate also stimulates the activation of pyruvate kinase. - Once we get to pyruvate, it all depends on how much oxygen is present. If there is enough oxygen (e.g. jogging), pyruvate will go through TCA, if there is low oxygen levels (e.g. sprint), fermentation and formation of lactate will take place. 12 Monday, January 18, 2016 Week 3 Lecture 6. Glycogen metabolism . - Berg et al., pp. 489‐492; 615‐635; Lodish et al., pp. 699‐703. - The “ Warburg” effect is a phenomenon concerning biological oxidation. Rapidly growing tumours metabolize glucose to lactate even when ample oxygen is present because tumours grow faster than blood vessels that feed them: environment becomes oxygen poor – hypoxia. Hypoxia activates HIF transcription factor. Active HIF switches on genes encoding glycolytic enzymes. Active HIF increase expression of VEGF – stimulates blood vessel formation. - Gluconeogenesis is somewhat the opposite of glycolysis; non-carbohydrate precursors (lactate, some amino acids, glycerol) are converted into glucose. This pathway takes place mainly in the liver and is important during extended exercise and fasting in order to keep blood glucose levels up. The brain is dependent on glucose, it doesn’t take any other form of energy! - Gluconeogeneis is not simply the reverse of glycolysis, not all the steps in the pathway are reversible. The picture below boxed the irreversible steps. 1 Monday, January 18, 2016 - In fact, in the gluconeogenesis pathway, the three irreversible steps of glycolysis are replaced by four other steps. • 1. Pyruvate —> Oxaloacetate (OAA) • 2. OAA —> PEP • 3. F-1,6-BP —> F-6P • 4. 6-6P —> glucose • Some amino acids and lactate —> pyruvate • Other amino acids —> OAA • Glycerol (from lipids) —> DHAP - Pyruvate carboxylase adds a carboxyl group to pyruvate and converts Pyruvate to OAA. This enzyme is located in the mitochondria, it contains biotin necessary for carboxylation and it required acetyl CoA although it is not a substate. - PEP carboxykinase: OAA+GTP—>PEP+GDP+CO2. PEP carboxykinase is cytosolic (contained in the cytosol) In order to convert pyruvate to PEP, there is first a carboxylation and followed by a decarboxylation. 2 Monday, January 18, 2016 - Fructose 1,6 Bisphosphatase converts F-1,6-BP —> F-6P + P. It is inhibiied by AMP and F-2,6-BP but activated by citrate. It is important that fructose 1,6 bisphosphatase and PFK aren’t operating simultaneously since it will burn ATP for absolutely no reason. - Fructose-6-phosphate is easily converted into glucose-6-phosphate (reversible step). Glucose-6-phosphatase then converts G-6P —> G + P. Glucose-6-ihosphatase is only found in tissues (liver, kidney) that regulate blood glucose levels. It is important to regulate free glucose levels. Remember that glucose can pass through the cell membrane but not the phosphorylated form. - Conditions that favour glycolysis inhibit gluconeogenesis and vice versa. Glycolysis is catabolic (breakdown) and energy yielding while gluconeogensis is anabolic (build- up) and energy consuming. - During intensive exercise, pyruvate made faster in muscle than citric acid cycle can oxidize it —> lactate production. Lactate in blood is then taken up by liver. The lactate converted back into glucose in liver through gluconeogenesis. The picture above is a simple illustration and the picture on the right is more complete. Important to know that the cardiac muscle is very rich in mitochondria. 3 Monday, January 18, 2016 - Glucose is stored as glycogen. The precursor of glycogen is uridine-diphosphate glucose. Breaking glycogen down, requires an inorganic phosphate group. 4 Monday, January 18, 2016 - Blood glucose must be tightly regulated. Both glucagon and insulin are hormones that regulate it. • Insulin released from the pancreas when blood glucose is high, stimulates synthesis of glycogen from glucose. • Glucagon released from pancreas when blood glucose is low, stimulates the breakdown of glycogen to glucose. • M u s c u l a r a c t i v i t y o r i t s anticipation stimulates release of epinephrine (adrenalin) from adrenal medulla. - Glucagon binding to receptor on liver cell, or epinephrine binding on muscle cell activates adenylate cyclase: ATP —> cAMP + PP. i 5 Monday, January 18, 2016 - READ BOOK! - Elevated cAMP levels activate protein kinase A (PKA). 6 Monday, January 18, 2016 Lecture 7. The tricarboxylic acid (TCA) cycle . - Berg et al., pp. 497‐516. - In order to regenerate NAD when there is no oxygen, our body undergoes lactic acid fermentation. But pyruvate can be oxidized if oxygen is present. - Pyruvate has to be first converted into carbon dioxide and acetyl CoA to enter the tricarboxylic acid cycle. • Carbohydrates —> pyruvate —> acetyl CoA • This pathway allows recovery of much greater portion of free energy than glycolysis and occurs in the mitochondria. • The mitochondria has two membranes (outer and inner), there is an inter membrane space in between them. The inner membrane encloses the mitochondrial matrix where the Kreb’s cycle takes place. - Acetyl CoA has a sulfhydryl group that binds (thioester bond) to the acetyl groups and that bond stores a lot of energy. The acetyl group of acetyl CoA completely oxidizes to two CO . • 2 • The 8 electrons from acetyl are transferred to NAD+ and FAD. - NADH and FADH forme2 during cycle reoxidized to NAD+ and FAD through action of respiratory or electron transport chain. - A proton gradient is created within the intermembrane of the mitochondria with the electrons from NADH and FADH and 2 transferring the low energy electrons to oxygen releasing water. Then, the gradient could be discharged through oxidative phosphorylation when protons flow back and pass by ATP synthase. 7 Monday, January 18, 2016 - The first step after glycolysis, if oxygen is present, is to oxidize pyruvate in the mitochondria. It is an irreversible reaction and is catalyzed by pyruvate dehydrogenase complex. The complex has over three distinct enzymes associated in a huge protein/coenzyme complex (60 polypeptide chains) The three distinct coenzymes derived from vitamins and they are E1 (thiamine • pyrophosphate), E2 (lipoic acid) and E3 (FAD). • Pyruvate + CoA + NAD+ —> Acetyl CoA + CO + NADH + H 2 + - Because the pyruvate dehydrogenase reaction is irreversible, it is highly regulated. • It is allosterically inhibited by end products (NADH and acetyl CoA). • It is switched off by phosphorylation; increased NADH/NAD+ ratios, acetyl CoA/ CoA ratios or ATP/ADP ratios promotes phosphorylation. • An accumulation of substrates (pyruvate and ADP) promotes dephosphorylation. - In the next step, TCA cycle begins (overall picture in following pages). 1. Acetyl CoA binds to oxaloacetate (4 carbon compound) forming citrate (citric acid) which is a tricarboxylic acid (3 carboxyl groups) with the enzyme citrate synthase. It is the hydrolysis of the thirster bond that releases the energy used to drive the reaction. Therefore, the equilibrium lyes far to the citrate formation. 2. The enzyme aconitase removes a water molecule and re-adds it forming an isomer called isocitrate; the hydroxyl has been moved. 8 Monday, January 18, 2016 3. Isocitrate dehydrogenase then oxidizes isocitrate’s hydroxyl group into a carbonyl. A NAD+ is also changed into NADH. Then, a carboxyl group is released forming alpha-ketoglutarate. This type of process is called an oxidative decarboxylation. In the next step, we go from alpha-ketoglutare (5 carbon dicarboxylic acid) to 4. succinate (4 carbon dicarboxylic acid). alpha-ketoglutarate + GDP + Pi —> succinate + GTP. 1. Alpha-ketoglutarate dehydrogenase removes a terminal carboxyl and forms a thioester with CoA. A NADH is produced. This reaction has the same mechanism as pyruvate dehydrogenase. This step is inhibited by NADH and succinyl CoA. 2. Succinyl CoA synthetase hydrolyses the thioester bond to release CoA. There are different form of this enzyme depending where it is. At the same time, the energy is captured to phosphorylate GDP —> GTP or ADP —> ATP. depending where the enzyme is located once again. 9 Monday, January 18, 2016 5. Since it is a cycle, succinate has to be reconverted into oxaloacetate. 1. Succinate dehydrogenase located in the inner membrane converts it into fumarate and electrons go to form a FADH . 2 2. Fumerase then adds a water across the double bond converting fumarate to malate. 3. Malate dehydrogenase completes the cycle but equilibrium favours malate. therefore it is not a very efficient reaction. 10 Monday, January 18, 2016 - The first reaction of the cycle lyes far to citrate (very negative free energy change) whereas the last reaction of the cycle lyes far to the malate (very positive free energy change). Therefore, the concentration of oxaloacetate remains low within the mitochondria matrix. In certain cases, oxaloacetate’s concentration can be so low that it’ll limit the rate of TCA. - Both decarboxylation steps (3 &4) have negative free energy change, hence the overall standard free energy change is low for the cycle. - Succinate dehydrogenase, unlike the other proteins, this enzyme is part of a respiratory complex of the inner mitochondrial membrane, it does not rely in the matrix but in the membrane. + AcetylCoA+3NAD +FAD+GDP+P → CoA+2CO +i ADH+FADH +GTP 2 2 11 Monday, January 18, 2016 Lecture 8. Regulation of the TCA. - Berg et al., pp. 516‐521; 639‐648. - Isocitrate dehydrogenase’s activity is important, high change in free energy. It is reduced by ATP and NADH and activated by ADP and NAD+. - Alpha-ketoglutarate dehydrogenase complex is also inhibited by ATP and NADH and Succinyl CoA. - The availability of oxaloacetate is the limiting factor of this cycle. - The intermediate molecules part of the cycle are very important and many are precursors of formation of other biomolecules. For example, citrate can be exported out of the mitochondria into the cytoplasm and used to synthesize fatty acids. 12 Monday, January 18, 2016 - Fatty acid oxidation can occur in mitochondria or peroxisomes. • Fatty acids —> Fatty Acyl CoA, it gets “activated”. This step uses ATP and occurs within the cytoplasm. • Pathways are similar in both compartments (mitochondria and peroxisome). • Products are Fatty Acyl CoA - 2 carbon + Acetyl CoA. 13 Monday, January 18, 2016 • In mitochondria, FADH 2 and NADH are oxidized by respiratory chain: Acetyl CoA enters TCA where 2 CO 2 are leaving. • In peroxisomes, FADH red2ces O toH O a2d acet2l2 CoA gets exported. - Ketone bodies are produced by the liver from fatty acids during periods of low food intake. In other words, they exist when gluconeogeneis and fatty oxidation occur simultaneously, insufficient oxaloacetate to react with acetyl CoA produced. - These ketone bodies are converted into acetyl-CoA which then enters the citric acid cycle and is oxidized in the mitochondria for energy. 14 Monday, January 25, 2016 Week 4 Lecture 9. Metabolism and physiology . - Lodish et al., 473‐503. - When gluconeogeneis (catabolic pathway) and fatty acid oxidation (anabolic pathway) occur simultaneously, there isn’t enough oxaloacetate to react with acetyl CoA. Therefore, ketone bodies are produced. - A consequence is the fact that ketone bodies are relatively strong acids and will lower the pH in the blood. Dangerous is pH is too low in the brain. • Oxaloacetate is an intermediate in gluconeogenesis. • When acetyl CoA concentration is too high in the mitochondrial matrix, it can react with itself and can form betahydroxybutyrate and acetoacetate. • This occurs in the liver and it will release these compounds in the blood stream. - This situation happens when one is fasting. • The body prefers carbohydrates as a source of energy. If you don’t eat all day, the stored glycogen is used up but the body needs to keep glucose in the blood for the brain to function. • In absence of dietary carbohydrates glycolysis limited and fats are going in the liver to extract energy. • Since glucose is needed, liver oxaloacetates are directed to the formation of glucose. • Some dietary amino acids converted into glucose by liver (gluconeogenesis) while other dietary amino acids are converted to acetyl CoA. 1 Monday, January 25, 2016 • Hence, there is an increase in the reliance of fats as energy source — breakdown of adipose tissue. Therefore, fatty acid oxidation and gluconeogenesis are occurring at same time . • Ketone bodies are produced by liver and exported in the bloodstream • In extreme conditions (starvation) brain adapts by allowing ketone bodies across blood brain barrier. - Ketone bodies are also present when one has diabetes. • There is a lot of glucose in the blood but cells aren’t able to take it all up because no insulin is present. Thus, no glycolysis going on in the tissues; there is high blood glucose but it cannot get into the cells. • The liver oxaloacetate is hence directed to the formation of glucose. • Some dietary amino acids converted into glucose by liver (gluconeogenesis) while other dietary amino acids are converted to acetyl CoA. • Therefore lipids are broken down and released into the bloodstream. All the fatty acids are broken down into acetyl CoA and ketone bodies. • The brain once again adapts by allowing ketone bodies to cross the blood brain barrier. - Both carbohydrates and fats can be used provide energy but carbohydrates are preferred; • Glucose is only source of energy for the brain since there is a blood/brain barrier. • Red blood cells have no mitochondria so have to rely on glycolysis. • High intensity exercise (sprint) driven almost exclusively by anaerobic glycolysis. The maximum human sprint distance (full speed) is about 200 meters and therefore pacing is critical for longer distance races. - Scenario of 800m race. He starts fast, he slows down, he feels good, he’s hanging on towards the end and can barely walk after the race. What happened? • Lactic acid build up early. Slow down in pace allowed for more aerobic metabolism in mid-race. • Last 100m: need for muscle ATP acute but lactic acid build up leads to increase in muscle proton concentration which inhibits PFK. Glycolysis stops and muscles stop working. 2 Monday, January 25, 2016 - Scenario of marathon: about 75% of calories come from carbohydrate reserves and 25% from fats. • Maximum carbohydrate energy storage is around 2000 calories (good for 20 miles) • As the race goes on, carbohydrates becomes depleted. If carbohydrate gets depleted before race finishes you hit the wall. • Fatty acids cannot supply the energy without carbohydrates since oxaloacetate limits the tricarboxylic acid cycle. The body has to keep doing gluconeogenesis to replenish oxalocatate but the brain and red blood cells need glucose. Hence, there is no way to provide muscles with adequate ATP to maintain pace. - The Atkins diet is basically cutting off carbohydrates forcing the body to use stored fat as energy source. • The proteins provide a source of oxaloacetate and some glucose through gluconeogenesis. • A downside of this is bad breath because ketone bodies are produced. • It is a state of constant ketosis which is not good longterm since ketone bodies do acidify the blood. 3 Monday, January 25, 2016 - Phospholipids consist of a glycerol which has 3 hydroxyl groups. Two of those groups are attached to two fatty acid chains via ester linkage and a third terminal one is attached to a phosphate (phosphatidic acid, PA). - Phosphates are usually linked to another group. For example, choline in phosphatidyl choline, PI. - Glycerol phosphate portion constitutes polar head, water loving. - Phospholipids are the major lipid component of cell membranes - The amphipathic phospholipids instantly form a phospholipid bilayer. These bilayers are permeable to small non-polar molecules e.g. CO2 and O2. They are partly permeable to water and impermeable to charged molecules, ions and larger uncharged molecules such as glucose. - Cellular membranes consist of proteins embedded in a phospholipid bilayer. - Most substances require special transport protein to pass across membranes. 4 Monday, January 25, 2016 Lecture 10. Metabolic integration. - Berg et al., pp. 654‐66; 791‐810. - Movement left to right is energetically favourable; it is entropy driven. The opposite from right to left is unfavourable. • To move a molecule against gradient (uphill) we must expend energy: G > 0 • Movement down a concentration (downhill) gradient releases energy: G < 0 - Passive diffusion is a downhill movement of a substance across a lipid bilayer. • Smaller non-polar or mildly polar molecules can diffuse across cell membranes. • The rate of passage across membrane is determined by concentration gradient, permeability coefficient; the more hydrophobic, the higher permeability coefficient. • For example: benzene (non polar) will diffuse faster than ethanol which is faster than ammonia or alanine (charged on both sides of the amino acid body even at neutral pH). - There are different types of membrane transport proteins: most substances require membrane transport protein to enter or leave the cell. • ATPase pumps — active transport uphill coupled to ATP hydrolysis. • Ion channels and uniporters — transport downhill such as facilitated diffusion. • Symporters and antiporters — uphill movement of one molecule coupled to downhill movement of another. - GLUT1 is an important protein (for red blood cells specifically). It has a glucose binding site, the binding leads to a conformational change flipping the channel towards the interior of the cell. Then the glucose is phosphorylated and gets trapped. 5 Monday, January 25, 2016 - Transporters have something similar to a K m GLUT1 (see picture on the right). The normal concentration of glucose in the blood is around 5 mM; hence, GLUT1 is nearly operating at nearly 75% of its maximum capacity. GLUT3 is very similar to GLUT1, it is found in neuronal tissue. - GLUT2 is present in the liver but is different than GLUT1 and GLUT3. GLUT2 in the liver only starts to take up glucose when the concentrations are very high after a meal for example. This enzyme has a relatively high K malue and it can also work in reverse and pump glucose out of the liver when blood glucose levels are low. - The movement is ions across membranes is more challenging than an uncharged molecule; it has to move against both a charge and a concentration gradient which might even be opposed at time. Z is the charge, F is the Faraday constant and V is the potential. - In the opposite direction, log C 2C 1ill give a number smaller than zero and yielding an negative free energy. - Cells in general maintain a potential gradient that is negative on the inside. Potassium concentrations are relatively high within the cell and sodium concentrations are relatively low. 6 Monday, January 25, 2016 - Three ion specific channels are illustrated on the right. a) doesn’t allow anything to pass through, b) + only allows the flow of Na ion and c) only allow potassium ions. Therefore, ions move down the concentration gradient and stops when the energy term from charge change (ZFV) is equal to the term from concentration change (2.3 RT log C /C ). 2 1 - The picture above illustrates that the entry of sodium into the cell is energetically favourable. 7 Monday, January 25, 2016 - There are many ATP-powered ion pumps divided in a few classes (P-class pumps, V- class proton pumps, F-class proton pumps & ABC superfamily). - P-class pumps transports only ions and there are 2 alpha (usually) and 2 beta subunits, it can exist as tetramers. It is the alpha subunit is involved in transport and ATP hydrolysis. + • An example is the animal Na / K ATPase (sodium potassium pump). It needs to keep potassium concentrations high within the cell to synthesize proteins and sodium concentrations low. For each ATP hydrolyzed, it pumps 3 Na +out and 2 K in. It is also inhibited by oubain and digoxin. 8 Monday, January 25, 2016 - V-class H pumps are similar in structure to the F-class proton pumps which are involved in oxidative phosphorylation and photosynthesis. • They pump protons into lysosomes and vacuoles. Enzyme is not phosphorylated during reaction cycle. • In order to compensate for the charge differential, these pumps are coupled to a chloride ion channel for effective acidification of the lumen. - ABC transporters (ATP Binding Cassette) have two transmembrane and two ATP binding domains. • These proteins are very common in bacteria. For example, bacteria permeases allow entry of key nutrients such as some amino acids. • An example is mammalian MDR (multiple drug resistance) transporters: it transports drugs to the exterior of the cell and the membrane orientation of the protein changes. • Another example is the CFTR protein: cystic fibrosis transmembrane regulator is a chloride channel protein in lungs and other tissues. 9 Monday, January 25, 2016 Lecture 11. Biological redox reactions (oxidative phosphorylation) . - Berg et al., pp. 525‐531. - NADH and FADH are f2rmed during cycle re- oxidized to NAD and FAD through action of respiratory or electron transport chain. -Respiratory chain transfers electrons to O 2o form H O.2Coupled to + formation of H gradient across inner membrane. - The discharge of gradient is coupled to ATP synthesis (oxidative phosphorylation) - NADH and Succinate come in and electrons are transferred to lots of electron carriers. - Redox reactions require an electron donor and electron acceptor (NAD and FAD). -In the electron transfer chain, each transfer from carrier to carrier is a redox reaction. Succinate + FAD —> fumarate + FADH . 2 - It can be considered as two half reactions. Succinate —> fumarate + 2e- + 2H (oxidation) • • FAD + 2e- + 2H —>+FADH (reduc2ion) 10 Monday, January 25, 2016 - The next reaction in the chain involves a non-heme ion (NHI). FADH + 2Fe 3+ (NHI) —> FAD + 2Fe 2+ (NHI) + 2H + - Therefore, oxidation is a gain of H/H character or a loss of elect
More Less

Related notes for BIOL 201

Log In


Join OneClass

Access over 10 million pages of study
documents for 1.3 million courses.

Sign up

Join to view


By registering, I agree to the Terms and Privacy Policies
Already have an account?
Just a few more details

So we can recommend you notes for your school.

Reset Password

Please enter below the email address you registered with and we will send you a link to reset your password.

Add your courses

Get notes from the top students in your class.