Biology 123: Week 4

Mitochondrial cristae contain the ETC and the ATP synthase complex.

NADH and FADH2 carry electrons to ETC. Carriers pump hydrogen ions - H+ gradient Chemiosmosis As H+ flow from down concentration gradient, ATP synthase synthesizes ATP from ADP + P.

Preparatory (prep) reaction

Occurs before the Krebs cycle 3-carbon pyruvate is converted to a 2-carbon acetyl group as CO2 is given off.

Citric acid cycle

Occurs in the mitochondrial matrix Consists of a series of reactions that return to their starting point (citric acid) Produces a lot of the NADH and all of the FADH2

Electron transport chain (ETC)

Located in the cristae of the mitochondria Series of carriers that pass electrons Electrons lose energy as they are passed along Cytochrome molecules are members O2 is final electron acceptor Forms water Without O2, ETC does not function.

Glycolysis

Glycolysis is the first step in the breakdown of glucose to extract energy for cellular metabolism. Nearly all living organisms carry out glycolysis as part of their metabolism. The process does not use oxygen and is therefore anaerobic (processes that use oxygen are called aerobic). Glycolysis takes place in the cytoplasm of both prokaryotic and eukaryotic cells. Glucose enters heterotrophic cells in two ways. Through secondary active transport in which the transport takes place against the glucose concentration gradient. Through a group of integral proteins called GLUT proteins, also known as glucose transporter proteins. These transporters assist in the facilitated diffusion of glucose. Glycolysis begins with the six carbon ring-shaped structure of a single glucose molecule and ends with two molecules of a three-carbon sugar called pyruvate (Figure 1). Glycolysis consists of ten steps divided into two distinct halves. The first half of the glycolysis is also known as the energy-requiring steps. This pathway traps the glucose molecule in the cell and uses energy to modify it so that the six-carbon sugar molecule can be split evenly into the two three-carbon molecules. The second half of glycolysis (also known as the energy-releasing steps) extracts energy from the molecules and stores it in the form of ATP and NADH, the reduced form of NAD. Step 1. The first step in glycolysis is catalyzed by hexokinase, an enzyme with broad specificity that catalyzes the phosphorylation of six-carbon sugars. Hexokinase phosphorylates glucose using ATP as the source of the phosphate, producing glucose-6-phosphate, a more reactive form of glucose. This reaction prevents the phosphorylated glucose molecule from continuing to interact with the GLUT proteins, and it can no longer leave the cell because the negatively charged phosphate will not allow it to cross the hydrophobic interior of the plasma membrane. Step 2. In the second step of glycolysis, an isomerase converts glucose-6-phosphate into one of its isomers, fructose-6-phosphate. An isomerase is an enzyme that catalyzes the conversion of a molecule into one of its isomers. This change from phosphoglucose to phosphofructose allows the eventual split of the sugar into two three-carbon molecules. Step 3. The third step is the phosphorylation of fructose-6-phosphate, catalyzed by the enzyme phosphofructokinase. A second ATP molecule donates a high-energy phosphate to fructose-6-phosphate, producing fructose-1,6-bisphosphate. In this pathway, phosphofructokinase is a rate-limiting enzyme. It is active when the concentration of ADP is high; it is less active when ADP levels are low and the concentration of ATP is high. Thus, if there is "sufficient" ATP in the system, the pathway slows down. This is a type of end product inhibition, since ATP is the end product of glucose catabolism. Step 4. The newly added high-energy phosphates further destabilize fructose-1,6-bisphosphate. The fourth step in glycolysis employs an enzyme, aldolase, to cleave 1,6-bisphosphate into two three-carbon isomers: dihydroxyacetone-phosphate and glyceraldehyde-3-phosphate. Step 5. In the fifth step, an isomerase transforms the dihydroxyacetone-phosphate into its isomer, glyceraldehyde-3-phosphate. Thus, the pathway will continue with two molecules of a single isomer. At this point in the pathway, there is a net investment of energy from two ATP molecules in the breakdown of one glucose molecule. Second Half of Glycolysis (Energy-Releasing Steps) So far, glycolysis has cost the cell two ATP molecules and produced two small, three-carbon sugar molecules. Both of these molecules will proceed through the second half of the pathway, and sufficient energy will be extracted to pay back the two ATP molecules used as an initial investment and produce a profit for the cell of two additional ATP molecules and two even higher-energy NADH molecules. Step 6. The sixth step in glycolysis (Figure 3) oxidizes the sugar (glyceraldehyde-3-phosphate), extracting high-energy electrons, which are picked up by the electron carrier NAD+, producing NADH. The sugar is then phosphorylated by the addition of a second phosphate group, producing 1,3-bisphosphoglycerate. Note that the second phosphate group does not require another ATP molecule. Here again is a potential limiting factor for this pathway. The continuation of the reaction depends upon the availability of the oxidized form of the electron carrier, NAD+. Thus, NADH must be continuously oxidized back into NAD+ in order to keep this step going. If NAD+ is not available, the second half of glycolysis slows down or stops. If oxygen is available in the system, the NADH will be oxidized readily, though indirectly, and the high-energy electrons from the hydrogen released in this process will be used to produce ATP. In an environment without oxygen, an alternate pathway (fermentation) can provide the oxidation of NADH to NAD+. Step 7. In the seventh step, catalyzed by phosphoglycerate kinase (an enzyme named for the reverse reaction), 1,3-bisphosphoglycerate donates a high-energy phosphate to ADP, forming one molecule of ATP. (This is an example of substrate-level phosphorylation.) A carbonyl group on the 1,3-bisphosphoglycerate is oxidized to a carboxyl group, and 3-phosphoglycerate is formed. Step 8. In the eighth step, the remaining phosphate group in 3-phosphoglycerate moves from the third carbon to the second carbon, producing 2-phosphoglycerate (an isomer of 3-phosphoglycerate). The enzyme catalyzing this step is a mutase (a type of isomerase). Step 9. Enolase catalyzes the ninth step. This enzyme causes 2-phosphoglycerate to lose water from its structure; this is a dehydration reaction, resulting in the formation of a double bond that increases the potential energy in the remaining phosphate bond and produces phosphoenolpyruvate (PEP). Step 10. The last step in glycolysis is catalyzed by the enzyme pyruvate kinase (the enzyme in this case is named for the reverse reaction of pyruvate's conversion into PEP) and results in the production of a second ATP molecule by substrate-level phosphorylation and the compound pyruvic acid (or its salt form, pyruvate). Many enzymes in enzymatic pathways are named for the reverse reactions, since the enzyme can catalyze both forward and reverse reactions. Outcomes of Glycolysis Glycolysis starts with glucose and ends with two pyruvate molecules, a total of four ATP molecules and two molecules of NADH. Two ATP molecules were used in the first half of the pathway to prepare the six-carbon ring for cleavage, so the cell has a net gain of two ATP molecules and two NADH molecules for its use. If the cell cannot catabolize the pyruvate molecules further, it will harvest only two ATP molecules from one molecule of glucose. Mature mammalian red blood cells are not capable of aerobic respiration—the process in which organisms convert energy in the presence of oxygen—and glycolysis is their sole source of ATP. If glycolysis is interrupted, these cells lose their ability to maintain their sodium-potassium pumps, and eventually, they die. The last step in glycolysis will not occur if pyruvate kinase, the enzyme that catalyzes the formation of pyruvate, is not available in sufficient quantities. In this situation, the entire glycolysis pathway will proceed, but only two ATP molecules will be made in the second half. Thus, pyruvate kinase is a rate-limiting enzyme for glycolysis. IN SUMMARY: GLYCOLYSIS Glycolysis is the first pathway used in the breakdown of glucose to extract energy. It was probably one of the earliest metabolic pathways to evolve and is used by nearly all of the organisms on earth. Glycolysis consists of two parts: The first part prepares the six-carbon ring of glucose for cleavage into two three-carbon sugars. ATP is invested in the process during this half to energize the separation. The second half of glycolysis extracts ATP and high-energy electrons from hydrogen atoms and attaches them to NAD+. Two ATP molecules are invested in the first half and four ATP molecules are formed by substrate phosphorylation during the second half. This produces a net gain of two ATP and two NADH molecules for the cell. Figure 4 shows the entire process of glycolysis in one image:

Fermentation

Produces 2 ATP per glucose molecule when O2 is not available Pyruvate reduced to lactic acid or ethanol Benefit Provides a rapid burst of ATP without O2 Drawback Fermentation products are toxic to cells. Fermentation yields 2 ATP per glucose. Cellular respiration yields 36 or 38 ATP Fermenting yeasts leaven bread and produce alcohol. Fermenting bacteria produce acids used to make yogurt, sour cream, and cheese. Soy sauce is made by adding a mold and a combination of yeasts and fermenting bacteria to soybeans and wheat. Metabolic pool - substrates are entry points for degradation or synthesis of larger molecules Catabolism - reactions that break down molecules Anabolism - reactions that synthesize molecules The key to losing weight is to use more calories than you take in. Combine exercise with a sensible diet. Prolonged aerobic exercise burns fat. Breathing and heart rate increase during exercise in order to supply the muscles with adequate oxygen. Energy is released when carbohydrates are oxidized and used to produce ATP. Removal of hydrogen atoms occurs during glycolysis, the prep reaction, and the Krebs cycle. NADH and FADH2 carry electrons to the electron transport chain (ETC) to produce ATP. Oxygen serves as the final acceptor of electrons, and water is produced. Fermentation produces fewer ATP than cellular respiration, but does not require oxygen.

Glycolysis

Takes place in the cytoplasm outside the mitochondria No oxygen is required Glucose is split into two pyruvates. Divided into Energy-investment steps Energy-harvesting steps Substrate-level ATP synthesis Net gain of 2 ATP 2 NADH generated

Phases of cellular respiration:

1. Glycolysis Occurs in cytoplasm, produces pyruvate, does not require O2 2. Preparatory (prep) reaction In the matrix of the mitochondria 3. Citric acid cycle In the matrix of the mitochondria 4. Electron Transport Chain (ETC) Series of electron carriers in the cristae of the mitochondria Oxygen required as last acceptor of electrons

The Citric Acid Cycle

Glycolysis occurred in the cytoplasm of the cell. Now pyruvate travels into the mitochondria. This is where the citric acid cycle occurs. Remember the mitochondria produces much of your cells energy. The mitochondria has folded membranes, giving it a lot of surface area. Pyruvate is "chemically groomed" before entering the citric acid cycle. It is converted into Acetyl Co-A. This is the molecule that enters the citric acid cycle. You can see here that puruvate loses one carbon atom. This gives you one molecule of NADH. This will be used to make ATP later. Acetyl CoA then goes to the citric acid cy Occurs in the matrix of the mitochondria. Completely breaks down the two remaining pyruvate molecules into CO2 . When you breath out, 2/3 of the CO2 you get rid of comes from the Citric Acid Cycle. One Acetyl CoA enters the cycle. What is produced: One ATP molecule. Three NADH molecules. Two CO2 molecules. One FADH2 Since there are two pyruvate molecules, each that has been converted you Acetyl CoA, you have to have two citric acid cycles to completely break down sugar. So, in order to find out what is made, double these numbers! 2 ATP 6 NADH 4 CO2 2 FADH2 How much energy have we made so far? Glycolysis A net of 2 ATP. Citric Acid Cycle 2 NADH from the production of Acetyl CoA. 2 ATP 6 NADH The whole idea of breaking apart sugar is to get ATP. We have only made four so far. We need more!

Pyruvate oxidation

Introduction Among the four stages of cellular respiration, pyruvate oxidation is kind of the odd one out; it's relatively short in comparison to the extensive pathways of glycolysis or the citric acid cycle. But that doesn't make it unimportant! On the contrary, pyruvate oxidation is a key connector that links glycolysis to the rest of cellular respiration. Overview of pyruvate oxidation At the end of glycolysis, we have two pyruvate molecules that still contain lots of extractable energy. Pyruvate oxidation is the next step in capturing the remaining energy in the form of \text{ATP}ATPA, T, P, although no \text{ATP}ATPA, T, P is made directly during pyruvate oxidation.

Citric Acid Cycle (Krebs Cycle)

Like the conversion of pyruvate to acetyl CoA, the citric acid cycle takes place in the matrix of the mitochondria. Almost all of the enzymes of the citric acid cycle are soluble, with the single exception of the enzyme succinate dehydrogenase, which is embedded in the inner membrane of the mitochondrion. Unlike glycolysis, the citric acid cycle is a closed loop: the last part of the pathway regenerates the compound used in the first step. The eight steps of the cycle are a series of redox, dehydration, hydration, and decarboxylation reactions that produce two carbon dioxide molecules, one GTP/ATP, and reduced forms of NADH and FADH2. This is considered an aerobic pathway because the NADH and FADH2 produced must transfer their electrons to the next pathway in the system, which will use oxygen. If this transfer does not occur, the oxidation steps of the citric acid cycle also do not occur. Note that the citric acid cycle produces very little ATP directly and does not directly consume oxygen. Steps in the Citric Acid Cycle Step 1. The first step is a condensation step, combining the two-carbon acetyl group (from acetyl CoA) with a four-carbon oxaloacetate molecule to form a six-carbon molecule of citrate. CoA is bound to a sulfhydryl group (-SH) and diffuses away to eventually combine with another acetyl group. This step is irreversible because it is highly exergonic. The rate of this reaction is controlled by negative feedback and the amount of ATP available. If ATP levels increase, the rate of this reaction decreases. If ATP is in short supply, the rate increases. Step 2. Citrate loses one water molecule and gains another as citrate is converted into its isomer, isocitrate. Steps 3 and 4. In step three, isocitrate is oxidized, producing a five-carbon molecule, α-ketoglutarate, together with a molecule of CO2 and two electrons, which reduce NAD+ to NADH. This step is also regulated by negative feedback from ATP and NADH and by a positive effect of ADP. Steps three and four are both oxidation and decarboxylation steps, which release electrons that reduce NAD+ to NADH and release carboxyl groups that form CO2 molecules. α-Ketoglutarate is the product of step three, and a succinyl group is the product of step four. CoA binds the succinyl group to form succinyl CoA. The enzyme that catalyzes step four is regulated by feedback inhibition of ATP, succinyl CoA, and NADH. Step 5. A phosphate group is substituted for coenzyme A, and a high- energy bond is formed. This energy is used in substrate-level phosphorylation (during the conversion of the succinyl group to succinate) to form either guanine triphosphate (GTP) or ATP. There are two forms of the enzyme, called isoenzymes, for this step, depending upon the type of animal tissue in which they are found. One form is found in tissues that use large amounts of ATP, such as heart and skeletal muscle. This form produces ATP. The second form of the enzyme is found in tissues that have a high number of anabolic pathways, such as liver. This form produces GTP. GTP is energetically equivalent to ATP; however, its use is more restricted. In particular, protein synthesis primarily uses GTP. Step 6. Step six is a dehydration process that converts succinate into fumarate. Two hydrogen atoms are transferred to FAD, producing FADH2. The energy contained in the electrons of these atoms is insufficient to reduce NAD+ but adequate to reduce FAD. Unlike NADH, this carrier remains attached to the enzyme and transfers the electrons to the electron transport chain directly. This process is made possible by the localization of the enzyme catalyzing this step inside the inner membrane of the mitochondrion. Step 7. Water is added to fumarate during step seven, and malate is produced. The last step in the citric acid cycle regenerates oxaloacetate by oxidizing malate. Another molecule of NADH is produced. Products of the Citric Acid Cycle Two carbon atoms come into the citric acid cycle from each acetyl group, representing four out of the six carbons of one glucose molecule. Two carbon dioxide molecules are released on each turn of the cycle; however, these do not necessarily contain the most recently-added carbon atoms. The two acetyl carbon atoms will eventually be released on later turns of the cycle; thus, all six carbon atoms from the original glucose molecule are eventually incorporated into carbon dioxide. Each turn of the cycle forms three NADH molecules and one FADH2 molecule. These carriers will connect with the last portion of aerobic respiration to produce ATP molecules. One GTP or ATP is also made in each cycle. Several of the intermediate compounds in the citric acid cycle can be used in synthesizing non-essential amino acids; therefore, the cycle is amphibolic (both catabolic and anabolic). Breakdown of Pyruvate In order for pyruvate, the product of glycolysis, to enter the next pathway, it must undergo several changes to become acetyl Coenzyme A (acetyl CoA). Acetyl CoA is a molecule that is further converted to oxaloacetate, which enters the citric acid cycle (Krebs cycle). The conversion of pyruvate to acetyl CoA is a three-step process. Step 1. A carboxyl group is removed from pyruvate, releasing a molecule of carbon dioxide into the surrounding medium. (Note: carbon dioxide is one carbon attached to two oxygen atoms and is one of the major end products of cellular respiration. ) The result of this step is a two-carbon hydroxyethyl group bound to the enzyme pyruvate dehydrogenase; the lost carbon dioxide is the first of the six carbons from the original glucose molecule to be removed. This step proceeds twice for every molecule of glucose metabolized (remember: there are two pyruvate molecules produced at the end of glycolysis); thus, two of the six carbons will have been removed at the end of both of these steps. Step 2. The hydroxyethyl group is oxidized to an acetyl group, and the electrons are picked up by NAD+, forming NADH (the reduced form of NAD+). The high- energy electrons from NADH will be used later by the cell to generate ATP for energy. Step 3. The enzyme-bound acetyl group is transferred to CoA, producing a molecule of acetyl CoA. This molecule of acetyl CoA is then further converted to be used in the next pathway of metabolism, the citric acid cycle. The citric acid cycle, shown in —also known as the tricarboxylic acid cycle (TCA cycle) or the Krebs cycle—is a series of chemical reactions used by all aerobic organisms to generate energy through the oxidation of acetate—derived from carbohydrates, fats, and proteins—into carbon dioxide. The cycle provides precursors including certain amino acids as well as the reducing agent NADH that is used in numerous biochemical reactions. Its central importance to many biochemical pathways suggests that it was one of the earliest established components of cellular metabolism; it may have originated abiogenically. The name of this metabolic pathway is derived from citric acid, a type of tricarboxylic acid that is first consumed and then regenerated by this sequence of reactions to complete the cycle. The cycle consumes acetate (in the form of acetyl-CoA) and water, reduces NAD+ to NADH, and produces carbon dioxide. The NADH generated by the TCA cycle is fed into the oxidative phosphorylation pathway. The net result of these two closely linked pathways is the oxidation of nutrients to produce usable energy in the form of ATP. Components of the TCA cycle were derived from anaerobic bacteria, and the TCA cycle itself may have evolved more than once. Theoretically there are several alternatives to the TCA cycle, however the TCA cycle appears to be the most efficient. If several alternatives independently evolved, they all rapidly converged to the TCA cycle. The citric acid cycle is a key component of the metabolic pathway by which all aerobic organisms generate energy. Through the catabolism of sugars, fats, and proteins, a two carbon organic product acetate in the form of acetyl-CoA is produced. Acetyl-CoA along with two equivalents of water (H2O) are consumed by the citric acid cycle, producing two equivalents of carbon dioxide (CO2) and one equivalent of HS-CoA. In addition, one complete turn of the cycle converts three equivalents of nicotinamide adenine dinucleotide (NAD+) into three equivalents of reduced NAD+ (NADH), one equivalent of ubiquinone (Q) into one equivalent of reduced ubiquinone (QH2), and one equivalent each of guanosine diphosphate (GDP) and inorganic phosphate (Pi) into one equivalent of guanosine triphosphate (GTP). The NADH and QH2 that is generated by the citric acid cycle is used by the oxidative phosphorylation pathway to generate energy-rich adenosine triphosphate (ATP). One of the primary sources of acetyl-CoA is sugars that are broken down by glycolysis to produce pyruvate that, in turn, is decarboxylated by the enzyme pyruvate dehydrogenase. This generates acetyl-CoA according to the following reaction scheme: CH3C(=O)C(=O)O- (pyruvate) + HSCoA + NAD+ → CH3C(=O)SCoA (acetyl-CoA) + NADH + H+ + CO2

Oxidative phosphorylation

Why do we need oxygen? You, like many other organisms, need oxygen to live. As you know if you've ever tried to hold your breath for too long, lack of oxygen can make you feel dizzy or even black out, and prolonged lack of oxygen can even cause death. But have you ever wondered why that's the case, or what exactly your body does with all that oxygen? As it turns out, the reason you need oxygen is so your cells can use this molecule during oxidative phosphorylation, the final stage of cellular respiration. Oxidative phosphorylation is made up of two closely connected components: the electron transport chain and chemiosmosis. In the electron transport chain, electrons are passed from one molecule to another, and energy released in these electron transfers is used to form an electrochemical gradient. In chemiosmosis, the energy stored in the gradient is used to make ATP. So, where does oxygen fit into this picture? Oxygen sits at the end of the electron transport chain, where it accepts electrons and picks up protons to form water. If oxygen isn't there to accept electrons (for instance, because a person is not breathing in enough oxygen), the electron transport chain will stop running, and ATP will no longer be produced by chemiosmosis. Without enough ATP, cells can't carry out the reactions they need to function, and, after a long enough period of time, may even die. In this article, we'll examine oxidative phosphorylation in depth, seeing how it provides most of the ready chemical energy (ATP) used by the cells in your body. The electron transport chain is a series of proteins and organic molecules found in the inner membrane of the mitochondria. Electrons are passed from one member of the transport chain to another in a series of redox reactions. Energy released in these reactions is captured as a proton gradient, which is then used to make ATP in a process called chemiosmosis. Together, the electron transport chain and chemiosmosis make up oxidative phosphorylation. The key steps of this process, shown in simplified form in the diagram above, include: Delivery of electrons by NADH and FADH_2 2 ​ start subscript, 2, end subscript. Reduced electron carriers (NADH and FADH_2 2 ​ start subscript, 2, end subscript) from other steps of cellular respiration transfer their electrons to molecules near the beginning of the transport chain. In the process, they turn back into NAD^+ + start superscript, plus, end superscript and FAD, which can be reused in other steps of cellular respiration. Electron transfer and proton pumping. As electrons are passed down the chain, they move from a higher to a lower energy level, releasing energy. Some of the energy is used to pump H^+ + start superscript, plus, end superscript ions, moving them out of the matrix and into the intermembrane space. This pumping establishes an electrochemical gradient. Splitting of oxygen to form water. At the end of the electron transport chain, electrons are transferred to molecular oxygen, which splits in half and takes up H^+ + start superscript, plus, end superscript to form water. Gradient-driven synthesis of ATP. As H^+ + start superscript, plus, end superscript ions flow down their gradient and back into the matrix, they pass through an enzyme called ATP synthase, which harnesses the flow of protons to synthesize ATP. We'll look more closely at both the electron transport chain and chemiosmosis in the sections below. The electron transport chain The electron transport chain is a collection of membrane-embedded proteins and organic molecules, most of them organized into four large complexes labeled I to IV. In eukaryotes, many copies of these molecules are found in the inner mitochondrial membrane. In prokaryotes, the electron transport chain components are found in the plasma membrane. As the electrons travel through the chain, they go from a higher to a lower energy level, moving from less electron-hungry to more electron-hungry molecules. Energy is released in these "downhill" electron transfers, and several of the protein complexes use the released energy to pump protons from the mitochondrial matrix to the intermembrane space, forming a proton gradient All of the electrons that enter the transport chain come from NADH and FADH_2 2 ​ start subscript, 2, end subscript molecules produced during earlier stages of cellular respiration: glycolysis, pyruvate oxidation, and the citric acid cycle. NADH is very good at donating electrons in redox reactions (that is, its electrons are at a high energy level), so it can transfer its electrons directly to complex I, turning back into NAD^+ + start superscript, plus, end superscript. As electrons move through complex I in a series of redox reactions, energy is released, and the complex uses this energy to pump protons from the matrix into the intermembrane space. FADH_2 2 ​ start subscript, 2, end subscript is not as good at donating electrons as NADH (that is, its electrons are at a lower energy level), so it cannot transfer its electrons to complex I. Instead, it feeds them into the transport chain through complex II, which does not pump protons across the membrane. Because of this "bypass," each FADH_2 2 ​ start subscript, 2, end subscript molecule causes fewer protons to be pumped (and contributes less to the proton gradient) than an NADH. Beyond the first two complexes, electrons from NADH and FADH_2 2 ​ start subscript, 2, end subscript travel exactly the same route. Both complex I and complex II pass their electrons to a small, mobile electron carrier called ubiquinone (Q), which is reduced to form QH_2 2 ​ start subscript, 2, end subscript and travels through the membrane, delivering the electrons to complex III. As electrons move through complex III, more H^+ + start superscript, plus, end superscript ions are pumped across the membrane, and the electrons are ultimately delivered to another mobile carrier called cytochrome C (cyt C). Cyt C carries the electrons to complex IV, where a final batch of H^+ + start superscript, plus, end superscript ions is pumped across the membrane. Complex IV passes the electrons to O_2 2 ​ start subscript, 2, end subscript, which splits into two oxygen atoms and accepts protons from the matrix to form water. Four electrons are required to reduce each molecule of O_2 2 ​ start subscript, 2, end subscript, and two water molecules are formed in the process. Overall, what does the electron transport chain do for the cell? It has two important functions: Regenerates electron carriers. NADH and FADH_2 2 ​ start subscript, 2, end subscript pass their electrons to the electron transport chain, turning back into NAD^+ + start superscript, plus, end superscript and FAD. This is important because the oxidized forms of these electron carriers are used in glycolysis and the citric acid cycle and must be available to keep these processes running. Makes a proton gradient. The transport chain builds a proton gradient across the inner mitochondrial membrane, with a higher concentration of H^+ + start superscript, plus, end superscript in the intermembrane space and a lower concentration in the matrix. This gradient represents a stored form of energy, and, as we'll see, it can be used to make ATP. Chemiosmosis Complexes I, III, and IV of the electron transport chain are proton pumps. As electrons move energetically downhill, the complexes capture the released energy and use it to pump H^+ + start superscript, plus, end superscript ions from the matrix to the intermembrane space. This pumping forms an electrochemical gradient across the inner mitochondrial membrane. The gradient is sometimes called the proton-motive force, and you can think of it as a form of stored energy, kind of like a battery. Like many other ions, protons can't pass directly through the phospholipid bilayer of the membrane because its core is too hydrophobic. Instead, H^+ + start superscript, plus, end superscript ions can move down their concentration gradient only with the help of channel proteins that form hydrophilic tunnels across the membrane. In the inner mitochondrial membrane, H^+ + start superscript, plus, end superscript ions have just one channel available: a membrane-spanning protein known as ATP synthase. Conceptually, ATP synthase is a lot like a turbine in a hydroelectric power plant. Instead of being turned by water, it's turned by the flow of H^+ + start superscript, plus, end superscript ions moving down their electrochemical gradient. As ATP synthase turns, it catalyzes the addition of a phosphate to ADP, capturing energy from the proton gradient as ATP. This process, in which energy from a proton gradient is used to make ATP, is called chemiosmosis. More broadly, chemiosmosis can refer to any process in which energy stored in a proton gradient is used to do work. Although chemiosmosis accounts for over 80% of ATP made during glucose breakdown in cellular respiration, it's not unique to cellular respiration. For instance, chemiosmosis is also involved in the light reactions of photosynthesis. What would happen to the energy stored in the proton gradient if it weren't used to synthesize ATP or do other cellular work? It would be released as heat, and interestingly enough, some types of cells deliberately use the proton gradient for heat generation rather than ATP synthesis. This might seem wasteful, but it's an important strategy for animals that need to keep warm. For instance, hibernating mammals (such as bears) have specialized cells known as brown fat cells. In the brown fat cells, uncoupling proteins are produced and inserted into the inner mitochondrial membrane. These proteins are simply channels that allow protons to pass from the intermembrane space to the matrix without traveling through ATP synthase. By providing an alternate route for protons to flow back into the matrix, the uncoupling proteins allow the energy of the gradient to be dissipated as heat. ATP yield How many ATP do we get per glucose in cellular respiration? If you look in different books, or ask different professors, you'll probably get slightly different answers. However, most current sources estimate that the maximum ATP yield for a molecule of glucose is around 30-32 ATP^{2,3,4} 2,3,4 start superscript, 2, comma, 3, comma, 4, end superscript. This range is lower than previous estimates because it accounts for the necessary transport of ADP into, and ATP out of, the mitochondrion. [More details] ^{5,6} start superscript, 5, comma, 6, end superscript Where does the figure of 30-32 ATP come from? Two net ATP are made in glycolysis, and another two ATP (or energetically equivalent GTP) are made in the citric acid cycle. Beyond those four, the remaining ATP all come from oxidative phosphorylation. Based on a lot of experimental work, it appears that four H^+ + start superscript, plus, end superscript ions must flow back into the matrix through ATP synthase to power the synthesis of one ATP molecule. When electrons from NADH move through the transport chain, about 10 H^+ + start superscript, plus, end superscript ions are pumped from the matrix to the intermembrane space, so each NADH yields about 2.5 ATP. Electrons from FADH_2 2 ​ start subscript, 2, end subscript, which enter the chain at a later stage, drive pumping of only 6 H^+ + start superscript, plus, end superscript, leading to production of about 1.5 ATP. One number in this table is still not precise: the ATP yield from NADH made in glycolysis. This is because glycolysis happens in the cytosol, and NADH can't cross the inner mitochondrial membrane to deliver its electrons to complex I. Instead, it must hand its electrons off to a molecular "shuttle system" that delivers them, through a series of steps, to the electron transport chain. Some cells of your body have a shuttle system that delivers electrons to the transport chain via FADH_2 start subscript, 2, end subscript. In this case, only 3 ATP are produced for the two NADH of glycolysis. Other cells of your body have a shuttle system that delivers the electrons via NADH, resulting in the production of 5 ATP. In bacteria, both glycolysis and the citric acid cycle happen in the cytosol, so no shuttle is needed and 5 ATP are produced. 30-32 ATP from the breakdown of one glucose molecule is a high-end estimate, and the real yield may be lower. For instance, some intermediates from cellular respiration may be siphoned off by the cell and used in other biosynthetic pathways, reducing the number of ATP produced. Cellular respiration is a nexus for many different metabolic pathways in the cell, forming a network that's larger than the glucose breakdown pathways alone