* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download THE CITRIC ACID CYCLE
Radical (chemistry) wikipedia , lookup
Mitochondrion wikipedia , lookup
Lactate dehydrogenase wikipedia , lookup
Electron transport chain wikipedia , lookup
NADH:ubiquinone oxidoreductase (H+-translocating) wikipedia , lookup
Amino acid synthesis wikipedia , lookup
Butyric acid wikipedia , lookup
Light-dependent reactions wikipedia , lookup
Photosynthesis wikipedia , lookup
Adenosine triphosphate wikipedia , lookup
Metalloprotein wikipedia , lookup
Biosynthesis wikipedia , lookup
Phosphorylation wikipedia , lookup
Basal metabolic rate wikipedia , lookup
Photosynthetic reaction centre wikipedia , lookup
Nicotinamide adenine dinucleotide wikipedia , lookup
Fatty acid synthesis wikipedia , lookup
Glyceroneogenesis wikipedia , lookup
Oxidative phosphorylation wikipedia , lookup
Evolution of metal ions in biological systems wikipedia , lookup
Microbial metabolism wikipedia , lookup
Fatty acid metabolism wikipedia , lookup
1 BI25M1 THE CITRIC ACID CYCLE LECTURE 1: AIMS: To review: the cycle in context, by considering how and why the use of oxygen to oxidise food molecules evolved. Lehninger: Instant Notes: Chapter 16 Section L1 2 1 LIFE EVOLVED 3.5-4.0 x 109 YEARS AGO. 0 First humans Extinction of dinosaurs Origin of reptiles Plants colonise land 500 Origin of multicellular organisms 1500 Oldest eukaryotic fossils 2500 Accumulation of atmospheric oxygen from photosynthetic cyanobacteria 3500 Oldest prokaryotic fossils 4500 Origin of Earth millions of years ago Early organisms obtained energy by taking up and breaking down simple molecules present in the environment. 3 They used this process: molecule simpler molecule (higher potential energy) (lower potential energy) energy ‘released’ The breakdown of the simple ‘food’ molecule is an exergonic reaction, (Energy Transformations lectures) and the organisms were able to use the ‘released’ energy. 2 THE LATER PART OF GLYCOLYSIS EVOLVED FIRST. Among the early, simple food molecules were ones similar to glyceraldehyde 3phosphate: CHO HCOH CH2OP 4 Some organisms managed to break this down into the chemically simpler pyruvate: COOC=O CH3 and ‘save’ some of the energy released as ATP, or a similar molecule. Thus evolved substrate-level phosphorylation (Energy Transformations lectures) and the later part of what we now call ‘glycolysis’ (Carbohydrates Lecture 4). and Intermediary Metabolism 5 3 EVOLUTION OF THE EARLY PART OF GLYCOLYSIS Simple food molecules were scarce. Some organisms evolved ways of making them from other, more complex molecules in the environment. Chief among these were hexoses, like glucose. glucose glyceraldehyde 3-phosphate pyruvate ATP Thus evolved the early, sugar-splitting (‘glyco-lysis’) part of glycolysis. 4 THE CHEMICAL NATURE OF GLYCOLYSIS Conversion of glucose (or glyceraldehyde 3-phosphate) to pyruvate is an oxidation. 6 The oxidation centres on the glyceraldehyde 3-phosphate dehydrogenase -catalysed reaction. In the same reaction, NAD+ is reduced to NADH + H+. hexoses glyceraldehyde 3-phosphate NAD+ pyruvate NADH + H+ Because of this, early organisms using the ‘energyproviding’ pathway faced a problem: when all the NAD+ is used up, flow stops. To let flow continue, another step, which regenerates NAD+, had to evolve. Some organisms developed this step: hexoses glyceraldehyde 3-phosphate NAD+ Others did this: pyruvate NADH + H+ NADH + H+ lactate NAD+ 7 hexoses glyceraldehyde 3-phosphate NAD+ pyruvate NADH + H+ ethanol NADH + H+ NAD+ Both mechanisms survive today, the former, for example, in vigorously exercising muscle, and when bacteria sour milk; the latter, for example, when yeast makes alcohol. Unlike hexoses pyruvate, NAD+ NADH + H+ which is a net oxidation, hexoses pyruvate NAD+ is not. NADH + H+ NADH + H+ lactate or ethanol NAD+ 8 The latter processes are ‘fermentations’ defined as ‘energy-yielding’ pathways in which no net change in the oxidation state of the reactants occurs. Because they involve no net oxidation, fermentations harvest only a small part of the potential energy of the glucose as ATP. (Just 2 mol ATP/mol glucose are generated Carbohydrates and Intermediary Metabolism Lecture 4). In summary: early organisms evolved ways of saving some of the potential energy of molecules in the environment using non-oxidative, catabolic fermentations. This involved making ATP by substratelevel phosphorylation. 9 5 MEANWHILE, OTHER ORGANISMS WERE USING SUNLIGHT. Molecules like hexoses were scarce, and organisms evolved several ways of generating them for themselves. All the ways worked like this: sunlight H2X + CO2 organic molecules X This is photosynthesis. For some organisms, ‘H2X’ was (and for some still is) H2S, or just H2. Others, including the cyanobacteria, used, as ‘H2X’, a molecule very abundant in many environments on Earth: H2O. 10 For such organisms, the released ‘X’ is, of course, O2. By about 2.5 x 109 years ago, their activity led to accumulation of O2 in the atmosphere to about present-day levels. 0 First humans Extinction of dinosaurs Origin of reptiles Plants colonise land 500 Origin of multicellular organisms 1500 Oldest eukaryotic fossils 2500 Accumulation of atmospheric oxygen from photosynthetic cyanobacteria 3500 Oldest prokaryotic fossils 4500 Origin of Earth millions of years ago 11 6 CONSEQUENCES OF OXYGEN ACCUMULATION Appearance in bulk of oxygen must have caused many problems. Oxygen oxidises, and disrupts the structure of the very organic molecules of which organisms are made. Many organisms must have become extinct. Others survived by evolving antioxidant defence mechanisms. Some actually managed to exploit the presence of the newly abundant molecule. 12 7 USE OF OXYGEN TO OXIDISE FOOD MOLECULES Glucose (C6H12O6) has many ‘high-energy’ electrons, so-called because they are part of electropositive H atoms. When the electrons associate with electronegative atoms, energy is released. (Energy Transformations Lecture 3). Some organisms evolved ways of ‘releasing’ and ‘saving’ this energy by removing H atoms from food molecules, and making the H atom electrons associate with electronegative NO3- or SO42-. Some present-day micro-organisms still do this. 13 Other organisms evolved a different method that was to become enormously successful. They made the H atom electrons associate with atoms of one of the most electronegative elements of all - the very material that had become abundant in their environment - oxygen. Their method involved stripping H atoms, two at a time, from what had originally been a food molecule, like glucose. The H atom electrons were passed to the oxidised form of a co-reactant, like NAD+ or FAD. (The reduced co-reactant in some cases is then used in an anabolic process useful to the organism (Energy Transformations Lecture 3)). 14 And the reduced co-reactant passed the H atom electrons on, through a series of redox carriers (the ‘terminal respiratory system’), until, eventually, they reached deliberately taken into the cell, and formed H2O. O2 This accomplished complete breakdown of the food molecule: glucose + 6O2 6H2O + 6CO2 Unlike the earlier fermentations, it really was an oxidation, and very exergonic. Much more of the potential energy of the food molecule was released, and could be ‘saved’ by making ATP. Thus evolved oxidative phosphorylation (Energy Transformations lectures). 15 Because they could harness so much of the potential energy of food molecules, organisms using this oxygen-involving process had an enormous evolutionary advantage. In addition, 16 8 OXYGEN USE WIDENED THE RANGE OF MOLECULES THAT COULD BE EXPLOITED AS FOOD. Some potential ‘foods’ have even more ‘high-energy’ electrons than hexoses, for example, fatty acids, e.g. CH3(CH2)14COO-. [Such molecules resemble petroleum hydrocarbons. These are also fuels because of their high content of H atom electrons (Energy Transformations Lecture 3).] The oxygen-involving apparatus that evolved allowed complete oxidation of hexoses like glucose (Section 7), but it also developed in such a way as to allow complete oxidation of fatty acids (and amino-acids) (Lipid Metabolism and How Organisms Handle Nitrogen lectures). So additional molecules became potential foods. 17 In summary: the primitive, non-oxygen-involving catabolism of hexoses survives, and is important today. For most cells, it is still the only way in which ATP can be made without oxygen (‘substrate-level phosphorylation’), and is used in eukaryotic cells lacking oxygen and/or the oxygen-involving ‘apparatus’ (Energy Transformations Lecture 3). [This is why glucose must be made (by gluconeogenesis), if glycogen stores are depleted and there is no dietary glucose (Carbohydrates and Intermediary Metabolism Lecture 5).] 18 9 WHAT IS THE OXYGENINVOLVING ‘APPARATUS’? In eukaryotic cells, the complete oxidation of food molecules occurs in mitochondria. (More primitive, partial catabolism of hexoses, by glycolysis, occurs in the cytosol.) H atoms are stripped from what were originally food molecules, and their electrons passed to NAD+ and FAD in a process called the ‘citric acid cycle’. Reduced NAD and FAD then pass the H atom electrons through a series of redox carriers (the ‘terminal respiratory system’), until they reach O2. Potential energy ‘released’ when they reach O2 is ‘saved’ by making ATP (‘oxidative phosphorylation’). (Section 7; Energy Transformations Lecture 3). 19 BI25M1 THE CITRIC ACID CYCLE LECTURE 2: AIMS: To review: the functions and processes of the: pyruvate dehydrogenase-catalysed reaction; citric acid cycle; glyoxylate cycle. Lehninger: Instant Notes: Chapter 16 Section L1 20 10 HOW DID CELLS BEGIN EXTRACTING FURTHER ENERGY FROM FOOD USING OXYGEN? Primitive, non-oxygen-involving catabolism of hexoses (‘glycolysis’) works like this (Sections 3,4): ATP hexose glyceraldehyde 3-phosphate NAD+ pyruvate NADH + H+ NADH+ + H+ lactate or ethanol NAD+ The last step (to lactate or ethanol) evolved to re-oxidise NADH made earlier in the process (Section 4). However, when O2 is being used, the last step isn’t needed, because the NADH can pass its H atom electrons along the terminal respiratory system to O2 (and so be re-oxidised). 21 In summary: some ATP has been made, by substrate-level phosphorylation, just as occurred in the absence of O2, but, additionally, when the lactate/ethanol step is avoided, the electrons from the two H atoms stripped from the food molecule and passed to NAD+, can now be passed (eventually) to O2, and more ATP can be made by oxidative phosphorylation. So, just avoiding the final step of the primitive process has begun the business of harvesting extra energy. 22 11 IF PYRUVATE ISN’T REDUCED TO LACTATE OR ETHANOL, WHAT HAPPENS TO IT? (Carbohydrates and Intermediary Metabolism Lecture 4) It is further oxidised. Another pair of ‘high-energy’-electroncontaining H atoms are stripped away, Again, their electrons are passed to NAD+. And, again, the electrons can pass to O2, and more ATP can be made by oxidative phosphorylation. The oxidation of pyruvate: is catalysed by pyruvate dehydrogenase; is complex; is essentially irreversible under intracellular conditions; occurs in mitochondria in eukaryotic cells. 23 This is what happens: Coenzyme A CO2 pyruvate COOC=O CH3 acetyl CoA NAD+ NADH + H+ CH3CSCoA O [Coenzyme A is a nucleotide-containing molecule that has an –SH group that can form a thio-ester link with carboxylic acids. (Lehninger, Edition 4 p.603; Edition 5 p.617] In summary: Further potential energy of the food molecule is ‘released’ and ‘saved’, as two more H atoms are stripped off and used to make ATP by oxidative phosphorylation. Notice the progressive break-down: glucose pyruvate 6C 3C acetyl CoA 2C [And remember glucose + 6O2 6H2O + 6CO2? This is the point where 2 of those 6 CO2 are made.] 24 12 WHAT HAPPENS TO THE ACETYL CoA? It is oxidised to CO2. The remaining H atoms are stripped off, two at a time. The electrons of most are passed to NAD+; some are passed to FAD. The oxidation of acetyl CoA; stripping away of the H atoms; reduction of NAD+ and FAD; all occur by a mechanism called the citric acid cycle (or tricarboxylic acid, TCA or Krebs’ cycle), which, in eukaryotic cells, occurs in mitochondria. Lehninger, Edition 4 p.607; Edition 5 p.621 Instant Notes, p.369 25 26 We can think of the cycle as a ‘black box’ (physicists talk about such things): it has to be there for the process to occur, but is itself unchanged during the process. Thus, flow through the cycle causes no net gain or loss of any of the 9 intermediates. The net changes that DO occur when one acetyl CoA is oxidised (i.e during one turn of the cycle) are: acetyl CoA 2CO2 3 (NADH + H+) 3H2O 1 FADH2 1 GTP (this is made from GDP, and is energetically equivalent to ATP) 27 A slightly more complicated version: acetyl CoA 2C oxaloacetate 4C 6C 4C 6C 4C 6C citrate NADH + H+ NADH + H+ CO2 FADH2 succinate 4C 4C 5C -ketoglutarate NADH + H+ CO2 GTP Yet another way of thinking about the cycle is: Attachment to carrier Carrier regenerated Breaking carrier where the ‘carrier’ is oxaloacetate. 28 The cycle is part of an oxygen-involving ‘apparatus’ concerned not just with complete oxidation of glucose, but also of fatty acids and amino-acids (Section 8). glucose fatty acids pyruvate oxidation acetyl CoA amino-acids CO2 (Lipid Metabolism and How Organisms Handle Nitrogen lectures). 29 13 THE CYCLE, AS A CYCLE, IS PURELY CATABOLIC. C flow through the cycle is: 2 C in (as acetyl group of acetyl CoA) 2 C out (as CO2), (Section 12) with no net production (or loss) of any of the 9 intermediates for possible use in anabolism. To emphasise this lack of an anabolic function, we can consider: 30 14 CARBOHYDRATE-FATTY ACID INTER-CONVERSION We know that: glucose pyruvate glycolysis acetyl CoA pyruvate dehydrogenase and that acetyl CoA can be catabolised through the citric acid cycle (Section 12). Acetyl CoA is also the building block of fatty acids (Lipid Metabolism lectures), so this process can also occur: glucose pyruvate acetyl CoA fatty acids fats Though this route, excess dietary carbohydrate is converted to fat and stored in adipose tissue. 31 15 IF GLUCOSE CAN BE CONVERTED TO FATTY ACIDS, IS THE REVERSE TRUE? How about this route? glucose pyruvate acetyl CoA fatty acids Well, it’s not possible, because the pyruvate-acetyl CoA reaction is irreversible (Section 11). How about this? Oxaloacetate is part of the gluconeogenesis pathway (Carbohydrates and Intermediary Metabolism Lecture 5) and ALSO one of the 9 citric acid cycle intermediates (Section 12). And fatty acids are catabolised to acetyl CoA (Lipid Metabolism lectures), 32 so could the fatty acids be converted into glucose like this? fatty acid acetyl CoA oxaloacetate glucose The answer is, again, no. There is no net flow of C from fatty acid to glucose by this route, because there is no net synthesis through the cycle of oxaloacetate (or any of the other cycle intermediates). The function of the cycle, remember, isn’t to make cycle intermediates, it’s to catabolise acetyl CoA to CO2 (Section 13). 33 Another way of looking at this is to remember what oxaloacetate is doing: it accepts the 2 C from acetyl CoA into the cycle; if it’s doing that, it can’t simultaneously be tapped off to make glucose. In summary: in many organisms (including all animal cells), fatty acids and acetyl CoA are not precursors of hexoses. (They were NOT among the short list of gluconeogenic precursors in Carbohydrates and Intermediary Metabolism Lecture 5). And the cycle, as a cycle, is purely catabolic. 34 Having said that parts of the cycle can form sections of linear metabolic pathways used in anabolism. For example, some amino-acids are metabolised to -ketoglutarate (one of the 9 intermediates), and through the route below, act as a source of glucose. glucose oxaloacetate some amino-acids -ketoglutarate part of the citric acid cycle 35 What happens is that the amino-acids ‘feed-in’ C, which replenishes oxaloacetate tapped off to make glucose, and part of the cycle is involved in this anabolic flow. Similar feed-in reactions enable other intermediates to be tapped off for anabolism. These ‘feed-in’ reactions are called anaplerotic (or replenishing) reactions. [How Organisms Handle Nitrogen Lecture 4] Because of this anabolic activity, the citric acid cycle is sometimes said to be amphibolic (i.e. to have catabolic and anabolic activity). But remember, as a cycle, it is purely catabolic. 36 16 THE GLYOXYLATE CYCLE This is evolution’s answer to the problem of converting fatty acids into carbohydrates. Some plant seeds store fat. At germination, the plant needs carbohydrates for many purposes, including building its structure. It lacks leaves, and so can’t make carbohydrates by photosynthesis. It needs to convert fatty acids from its fat stores into glucose. Similarly, some bacteria, protozoa and fungi grow on fatty acids with little or no carbohydrate present. They, too, must convert fatty acids into glucose. Such organisms have 2 enzymes, which animal cells lack. 37 These work with enzymes of part of the citric acid cycle, to produce another, anabolic, cycle: the glyoxylate cycle. C flow through the citric acid cycle is: (Section 12) 2Cin acetyl CoA 2 CO2 2Cout C flow through the glyoxylate cycle is: fatty acids 2Cin acetyl CoA part of the citric acid cycle *the 2 additional enzymes malate 4C isocitrate 6C * * glyoxylate 2C acetyl CoA 2Cin succinate 4Cout carbohydrates 38 or, more simply: 2Cin acetyl CoA succinate 4Cout acetyl CoA 2Cin In plant seeds, the two additional enzymes are in organelles called glyoxysomes.