Biochem Test 2

Energy metabolism can be defined as

"The pathways involved in the generation or storage of metabolic energy".

Therefore, hormones and signal molecules are also referred to as

'ligands'. Ligands bind to receptors. This is an image of a beta adrenergic receptor which is a transmembrane protein to which epinephrine binds. In this case, epinephrine is the ligand. Once the extracellular signal is received (by the receptor), the signal must be transduced to the inside of the cell. This is the next step in signal processing

The process of cell signaling can be broken down into three steps

1. Reception of the extracellular signal 2. Transduction of the signal from the outside of the cell to the inside of the cell 3. Cellular response to the signal

Two carbons (originating from acetylCoA) are added one at a time to an elongating fatty acyl chain. A few things are important here.

1.Where does that acetylCoA originate from? Recall that our bodies are in the mode of synthesizing fat when we have excess intake of carbohydrates. So when glucose is plentiful, it will first be used to replenish our body's glycogen stores, and then the excess will be broken down through glycolysis and the pyruvate dehydrogenase reaction to generate acetylCoA. When the acetylCoA is not required to generate energy, it won't be shuttled into the citric acid cycle, but rather, will be diverted to fat synthesis. 2.The other thing to consider is that anabolic processes are reductive processes. That is, we have to inject or donate electrons into the newly synthesized compound, in this case, fatty acids. Where do these electrons come from? They also come from reducing equivalents. And in the case of many biosynthetic reactions, NADPH is the reduced form of the reducing equivalent, NADP+ , which donates those electrons. This is a bit different from what we saw in gluconeogenesis, where NADH was the actual electron donor. Make sure to remember this difference. 3.Also, a tricarboxylate transport system must move acetylCoA (via a citrate intermediate) out of the mitochondrial matrix into the cytosol, because that is where fat synthesis takes place.

Fat Metabolism

A lot of attention has been paid to the "battle of the bulge" in recent decades, and fat has gotten a bad rap. In actuality, we get fat from consuming too many calories, whether from sugar, fat or protein. When food companies started to produce "low fat" or "non fat" versions of food, they sometimes added sugar to make the foods taste better. Fat is what actually makes food taste so darn good! So even though consumers were eating foods that were lower in fat, they were actually consuming just as many calories. So goes the old adage: "buyer beware"....

Synthesis Of ATP Through Oxidative Phosphorylation

ATP synthesis, or the oxidative phosphorylation of ADP to ATP, is catalyzed by complex V, which is also known as the F1-F0 ATP synthase. The F1-F0 ATP synthase is composed of multiple subunits. The F0 subunit is the proton channel which spans the inner mitochondrial membrane which is responsible for allowing protons to enter the matrix, while the F1 subunit is the bulbous portion of the complex on the matrix side of the inner membrane, which comprises the ATP synthase enzyme responsible for synthesizing ATP. The F0 and F1 subunits are held together by a protein 'stalk' which serves to connect the two subunits. The appearance of the F1-F0 ATP synthase has the appearance of a mushroom protruding into the matrix on the inside of the inner membrane.

Stage 2: AcetylCoA Enters The Citric Acid Cycle

AcetylCoA can now enter the citric acid cycle, which will complete the oxidation of glucose to carbon dioxide. Recall that acetylCoA is a two-carbon compound and it is broken down to two molecules of carbon dioxide containing one carbon each. The net overall reaction of the citric acid cycle is the production of three molecules of NADH, one molecule of FADH2, one molecule of GTP, two molecules of carbon dioxide, and free coenzyme A. There are eight enzymes in the citric acid cycle, and all of these are compartmentalized within the mitochondria, All citric acid cycle enzymes are soluble within the matrix, with the exception of the enzyme, succinate dehydrogenase, which is a membrane protein that is contained in the inner mitochondrial membrane.

Synthesis And Breakdown Pathways

AcetylCoA is also a precursor for cholesterol synthesis, and the synthesis of some amino acids! Also, the breakdown of some amino acids lead to the production of pyruvate or acetylCoA!

The Citric Acid Cycle: An Overview -- Movie Narrative

Aerobic respiration is the most important process used to produce energy for a cell. It involves several pathways including: glycolysis, pyruvate oxidation, the citric acid cycle, and electron transport. Various sugars, fats, and proteins enter these pathways, and they can all be broken down to produce ATP energy for a cell. In this animation, we will focus on the citric acid cycle--also called the tricarboxylic acid (TCA) cycle or Krebs cycle. The citric acid cycle takes place in the matrix, or fluid, of the mitochondrion. This is where mitochondrial DNA is found and where fatty acid breakdown takes place. The citric acid cycle involves eight chemical reactions that use acetyl CoA and oxaloacetate to produce carbon dioxide, NADH, ATP, and FADH2. The NADH and FADH2 are electron carriers that can be used in the electron transport chain to make more ATP for a cell. Let's take a look at the major steps of the citric acid cycle. In the first step of the cycle, a 2-carbon molecule and a 4-carbon molecule are combined to form a 6-carbon molecule. That 6-carbon molecule undergoes multiple biochemical changes during the cycle, and at the end the original 4-carbon molecule is produced. Each time a carbon molecule loses one carbon, a carbon dioxide is released. So, two carbon dioxides are formed during the conversion of the 6-carbon molecule back to the 4-carbon molecule. So, where does the 2-carbon molecule come from to begin the citric acid cycle? This molecule, acetyl CoA, is made from pyruvate. Pyruvate is a product of glycolysis, the other major ATP producing cycle in the cell. Pyruvate is transported into the mitochondrial matrix where it is oxidized to acetyl CoA by pyruvate dehydrogenase. This step produces one NADH and one carbon dioxide for each of the two pyruvate molecules made from glucose. The acetyl CoAs are the starting point for the citric acid cycle. In the citric acid cycle, the acetyl group from acetyl CoA is transferred to oxaloacetate to form citrate. Four different enzymatic reactions then lead to the formation of succinate. These steps produce two carbon dioxides, two NADHs, and one ATP. Succinate is then recycled back to oxaloacetate through three more reactions. These steps produce FADH2 and one more NADH. All of the energy molecules made in the citric acid cycle, along with those produced by glycolysis, are essential for fully functioning cells, as errors in these pathways can lead to lifethreatening diseases.

So it makes sense that we generate these two reducing equivalents in each round.

After a single round of beta oxidation, you have shortened your acyl group by two carbons. Then the shortened acylCoA starts the process all over again, entering a subsequent round of beta oxidation and cleaving two more carbons. This process repeats itself (yes, rinse and repeat) until all of the acyl group has been cut up into twocarbon acetylCoAs. That is, for even-numbered fatty acids. It is a bit of a different story for odd-numbered fatty acids, but the basic process is the same!

Values For Glycolysis Reactions

Again, glycolysis is a highly regulated pathway. Enzymes that catalyze reactions 1 and 3 are targets for the control of flux through glycolysis as these reactions are associated with a large change in free energy and as such are considered to be irreversible. These tend to be good points of regulation within a pathway.

Net Reaction of Pentose Phosphate Pathway

Alternatively, the carbons are further metabolized to two fructose-6-phosphate and one glyceraldehyde-3-phosphate from three original glucose-6-phosphate molecules. These intermediates can then feed into glycolysis and be further metabolized for energy generation.

Triacylglycerols Are Broken Down In Adipocytes

An enzyme called lipase releases the free fatty acids from the glycerol moiety by hydrolyzing the ester bonds

Anaplerosis: The replenishment Of Citric Acid Cycle Intermediates

Anaplerotic pathways for replenishment of citric acid cycle intermediates are shown with red arrows. In animals, the synthesis of oxaloacetate from pyruvate is quantitatively the most important anaplerotic reaction. Pyruvate carboxylase catalyzes this reaction.

Glycolysis Vs Gluconeogenesis

As mentioned earlier, gluconeogenesis is not simply the reverse of glycolysis. While it does share some of the same near-equilibrium enzymes, steps 1, 3 and 10, catalyzed by glucokinase, phosphofructokinase and pyruvate kinase must be by-passed as these reactions are irreversible. These three reactions are by-passed by glucose-6- phosphatase, fructose bisphosphatase, PEP carboxykinase and pyruvate carboxylase.

Protein As An Energy Source

As mentioned earlier, the breakdown of amino acids derived from protein can lead to the production of pyruvate or acetylCoA as well as to other intermediates in glycolysis or the citric acid cycle. This is how our body breaks down protein for energy.

Glycogen Synthase and Glycogen Phosphorylase Activity

As mentioned in the introduction to this module, some enzymatic reactions in metabolism are very tightly regulated and controlled. The reason for this, is that we want to ensure that futile cycling is not occurring, such as glycogen breakdown occurring simultaneously with glycogen synthesis. That would be a waste of energy. We want these pathways to proceed according to the metabolic circumstances of the organism. We also want to very tightly match the supply of energy, or ATP, with the demand for energy depending on metabolic needs. Yes, just like economics, it is about supply and demand. Not all enzymes in a pathway are regulated. We will focus on the key enzymes in metabolic pathways that regulate the overall flux through a given pathway. In this case, glycogen breakdown and synthesis are regulated by glycogen phosphorylase and glycogen synthase, respectively. These enzymes are controlled both by allosteric regulation and covalent regulation by phosphorylation

Fermentation

As we saw in the net reaction for glycolysis, glycolysis consumes NAD+ , reducing it to NADH. In order for glycolysis to continue to convert glucose to pyruvate, NAD+ must be regenerated from NADH. Under circumstances when O2 is present, this is done by the electron transport chain in the mitochondria. However, when O2 is absent, there are other reactions in the cell which can regenerate NAD+. This process, which occurs when O2 is absent is called fermentation. Fermentation is a pathway that produces ATP with no net oxidation of carbon

The Citric Acid Cycle

As you can see from the citric acid cycle, it begins and ends with oxaloacetate being consumed and regenerated. Note the number and type of reducing equivalents that are generated (3 NADH and 1 FADH2), GTP is generated (and that is also a form of energy that is equivalent to ATP), and lastly, two molecules of carbon dioxide are generated as well. Remember that this is stage 2 in the oxidation of pyruvate in the mitochondria. Therefore, we are drawing away electrons from the acetylCoA, and these are passed to the reducing equivalents NAD+ and FAD. Carbon dioxide is the waste product of metabolism that will eventually make its way out of the cell, into the bloodstream and exhaled from our lungs. Finally, coenzyme A is regenerated, because that is what coenzymes do! They are reused and recycled! So efficient! On the topic of coenzymes, how about all of the NADH and FADH2 reducing equivalents that we generated in glycolysis, the pyruvate dehydrogenase reaction and the citric acid cycle? Well, those need to be recycled and reused as well. We will see how those are regenerated through the electron transport chain...

Breakdown Of Fat From Stored Triacylglycerols

At rest or in between meals when glucose levels are low, the hormone glucagon is released and stimulates the lipase to breakdown fats to provide energy. Through the process of beta oxidation, the free fatty acids will be broken down and oxidized to generate the reducing equivalents, NADH and FADH2, as well as acetylCoA, which will feed into the citric acid cycle to generate some energy in the form of GTP and even more reducing equivalents. Do you remember how many NADH molecules and FADH2 molecules are generated from one round of the citric acid cycle?

In summary, metabolism can be broken down into two processes: catabolism and anabolism

Catabolism comprises processes in which complex biomolecules are broken down into simple molecules, whereas anabolism comprises processes in which complex biomolecules are being synthesized

Creatine Phosphate Buffers ATP Levels

Creatine phosphate is a high energy phosphate compound that serves as an energy buffer inside of the cell. It is readily available to buffer ATP levels as they drop during the onset of exercise or during very high intensity exercise. The enzyme creatine kinase catalyzes the transfer of a phosphate group from creatine phosphate to ADP to replenish ATP levels when they drop. Creatine is made in our body and consumed when we eat meat. Some athletes will consume a creatine supplement to augment their reserve of creatine phosphate. Some studies have shown that it can be an effective ergogenic aid for high intensity exercises like sprinting and weight lifting because it will give that extra boost of energy. But there are also studies that have shown it not to be effective. So the jury may be out on the use of creatine as an ergogenic aid. Nevertheless, it is still widely used, and not inexpensive. But why not just spend a little more time at the gym and keep your money in your wallet?

The breakdown of macronutrients is referred to as a "catabolic" process, or catabolism.

During catabolic processes, larger more complex macronutrients are broken down to smaller simple compounds like carbon dioxide. examples include glycolysis and citric acid cycle. The larger more complex molecule of glucose will be broken down through the pathway of glycolysis to yield a smaller compound called pyruvate. Pyruvate can then be further broken down to acetylCoA which can then be further broken down through the citric acid cycle pathway to carbon dioxide. Throughout the progression of the original glucose molecule through these pathways, energy, in the form of ATP, is generated.

Ketone Bodies

During conditions such as fasting or starvation, the body will draw on its carbohydrate stores located in the liver as glycogen. Remember that the brain prefers glucose as a fuel, and therefore after long periods of fasting, will use up all of the body's carbohydrate reserves. Now we are in trouble. Remember, that is why breakfast is important!! But wait, our body still has large reserves of fat in our adipose tissue. Of course, some of us have more or less than others... And that folks, is why you don't go on the television show "Survivor" to get stranded on an island for 39 days without building up your fat stores first! Sure, you want to be camera ready with a sleek physique, but you can be sure you may not do well surviving on eating cockroaches, rats and squirrels for very long without adequate fat stores in your adipose tissue! My apologies to this reference if you have not seen the show before! But anyways, hopefully you get the idea. Once our carbohydrate stores in the liver have been used up, we can take advantage of our fat stores because the liver has the ability to convert fats, via an acetylCoA intermediate, to ketone bodies. Our bodies are able to produce the following ketone bodies: acetone, acetoacetate, and beta-hydroxybutyrate.

Control of Flux Through Gluconeogenesis

Flux through gluconeogenesis is regulated reciprocally to flux through glycolysis to avoid futile cycling as mentioned earlier. A key point in the regulation of gluconeogenesis occurs at fructose bisphosphatase. In contrast to phosphofrutokinase which catalyzes the opposite reaction, fructose bisphosphatase is inhibited by high concentrations of AMP and fructose-2,6-bisphosphate.

Values for Glycolysis Reactions

Given the large changes in free energy that we saw in glycolysis, how can the reverse pathway of gluconeogenesis proceed?

One example of signal transduction is the mechanism of action of glucagon

Glucagon is the signal molecule that is released from the pancreas when blood glucose levels are low. Glucagon travels through the blood stream and subsequently binds to the glucagon receptor on liver cells to elicit the cellular response which is the release of glucose from its glycogen stores.

Gluconeogenesis

Gluconeogenesis is the de novo synthesis of glucose that occurs when the body's stores of glycogen are low. The brain needs and prefers glucose to continue to function. Gluconeogenesis takes place in the liver, and to a lesser extent in the kidneys. Precursors such as some amino acids, some citric acid cycle intermediates and lactate via pyruvate can be converted to oxaloacetate, which is subsequently converted to glucose through gluconeogenesis.

Overview Of The Breakdown Of Glucose To CO2

Glucose is gradually broken down in phases. The first phase of glucose metabolism is glycolysis and occurs in the cytosol. In glycolysis, glucose is broken down from a sixcarbon compound to two three-carbon compounds, called pyruvate. The net output of glycolysis is the generation of two molecules of NADH and two molecules of ATP. The oxidation of pyruvate occurs inside the mitochondrion and occurs in two stages. First, the conversion of the three-carbon pyruvate to a twocarbon molecule of acetylCoA, followed by the oxidation of acetylCoA to carbon dioxide through the citric acid cycle. The oxidation of pyruvate to acetylCoA is catalyzed by the enzyme, Pyruvate Dehydrogenase (PDH is the abbreviation for the name of this enzyme), which is located in the mitochondrion. The acetylCoA can then further be broken down through the citric acid cycle which is also located inside the mitochondrion. The citric acid cycle is a series of eight reactions which break down the two-carbon acetylCoA to the one-carbon molecule of CO2, while at the same time generating three molecules of NADH, one molecule of FADH2 and one molecule of GTP. All of the reducing equivalents, namely NADH and FADH2, will go on to the final stage of metabolism, which is the electron transport chain. Through the electron transport chain, the potential energy in the reducing equivalents will be converted to the chemical energy of ATP.

Glycogen Breakdown and Synthesis

Glycogen breakdown and glycogen synthesis are regulated by the enzymes glycogen phosphorylase and glycogen synthase, respectively. As you can see, glycogen phosphorylase catalyzes the phosphorolysis of glycogen with the addition of Pi. This is different than a hydrolysis reaction where water is added instead of Pi. The action of glycogen phosphorylase releases a glucose residue in the form of glucose-1-phosphate. The phosphoglucomutase enzyme converts this to glucose-6-phosphate, which can then enter glycolysis at reaction 2 as we will see shortly. Glycogen synthase catalyzes the synthesis of glycogen from glucose-1-phosphate, using UTP as energy to drive this reaction forward. Like ATP, UTP can be used as an energy source.

Recall that vertebrates store glucose in the form of glycogen

Glycogen is a highly branched structure with a single reducing end and several nonreducing ends. The alpha-1,4 bonds link the glucose subunits linearly, while the alpha1,6 bonds form the branch points. Glucose residues are sequentially removed from several non-reducing ends during glycogen degradation simultaneously, providing a rapid surge of glucose release when it is needed by the body.

The Citric Acid Cycle Is Amphibolic

Here is a fun fact: The citric acid cycle is amphibolic, which means that it is both catabolic and anabolic in nature. We know intuitively that the citric acid cycle is catabolic. We view its primary function as breaking down and oxidizing acetylCoA that originated from glucose or fat to generate ATP and reducing equivalents. But it also has another purpose, and that is to produce starting materials for the biosynthesis of some compounds. For example, citrate is siphoned off for the production of fatty acids and steroids, and succinylCoA is siphoned off for the synthesis of heme and chlorophyll. Alpha-ketoglutarate and oxaloacetate are siphoned off for the synthesis of some amino acids as well as purines and pyrimidines, respectively. But the cell cannot keep siphoning off citric acid cycle intermediates when they are required without replenishing them so that there will always be adequate amounts of oxaloacetate to begin the cycle again. This process of replenishing citric acid cycle intermediates is referred to as 'anaplerosis'.

Allosteric Regulation Of Glycogen Synthase And Glycogen Phosphorylase

Here is how glycogen synthase and glycogen phosphorylase are regulated allosterically. The dotted lines and arrows indicate allosteric regulation of each enzyme. Glycogen synthase is activated by high concentrations of glucose-6-phosphate. The high concentration of glucose-6-phosphate signals to the cell that there is plenty of carbohydrate available to be stored as glycogen for later use

Products Of Triacylglycerol Breakdown: Fate Of Fatty Acids

Here we see the fate of fatty acids as they exit the adipose tissue into the bloodstream. At rest, fatty acids are the preferred fuel for muscle. The liver will also use fat for energy. Once fatty acids are taken up inside the liver or muscle, they will be broken down by the process of beta oxidation. Before a fatty acid enters the pathway of beta oxidation, it must first be "activated".

Products Of Triacylglycerol Breakdown: Fate Of Glycerol

Here we see the fate of the glycerol that leaves the adipose tissue. It enters the bloodstream and is taken up by the liver, converted to glucose via gluconeogenesis and released back into the bloodstream where it can be taken up and used by tissues, such as the brain, for energy.

Net Reaction of Gluconeogenesis

Here we see the net reaction of gluconeogenesis beginning from pyruvate. We need to invest a total of 4 ATP plus 2 GTP (note: that is much more than the net of two ATP that we obtain from glycolysis). Remember that GTP can be used as an energy source, just like ATP. So, it is very costly to synthesize glucose from scratch! We also need to use 2 NADH. This makes sense as we are reducing pyruvate back to glucose. Then the glucose can be exported from the liver to meet the body's demand for glucose under conditions where the body has nearly used up its glycogen stores in between meals.

Covalent Regulation Of Glycogen Synthase And Glycogen Phosphorylase By Phosphorylation

Here, we see how glycogen synthase and glycogen phosphorylase are regulated covalently. Covalent addition of a phosphate group to an enzyme can act as a switch that turns the enzyme on or turn it off, depending on the enzyme in question. In the case of glycogen phosphorylase, phosphorylation by an enzyme called a kinase converts it to its active form. Removal of the phosphate group by an enzyme called a phosphatase converts it back to its inactive form. In contrast, phosphorylation of glycogen synthase converts synthase to its inactive form while removal of the phosphate group converts it to its active form. There are hormonal signals that regulate the phosphorylation and dephosphorylation by the kinase and phosphatase, respectively. The hormone insulin stimulates dephosphorylation by the phosphatase, while the hormone glucagon and epinephrine stimulate phosphorylation by the kinase

Beta Oxidation Of Fatty Acids In Matrix continued

If we take a 16 carbon fatty acid such as palmitate, and "activate" it to generate palmitoylCoA, 7 round of beta oxidation will break it down fully to 8 molecules of acetylCoA, 7 FADH2 and 7 NADH. Then what do we do with the products of beta oxidation? Well, the same thing that we do with the products of glycolysis and the pyruvate dehydrogenase reaction when we break down glucose. The acetylCoA is treated the same when we need to generate energy. It enters the citric acid cycle and is further oxidized and broken down to carbon dioxide! The reducing equivalents will also have the same fate. They will donate their electrons to the electron transport chain to generate ATP through oxidative phosphorylation. There is also an alternative fate for acetylCoA that occurs under conditions of starvation. That is, ketogenesis. The brain cannot use fatty acids for energy, but we can convert fatty acids to ketone bodies in the liver. And during fasting or under extreme circumstances such as starvation when glucose levels are low because the body's liver glycogen stores have been depleted, the brain can adapt to using ketone bodies as an energy source.

Fat Metabolism: Some Fats Are Healthy

In reality, there are healthy fats that our body needs, like omega-3 three fatty acids. Oily fish, like salmon, are an excellent source of omega-3s. Our body does not produce these types of fatty acids, and therefore they must be consumed in our diet as they are precursors for important compounds that help to regulate our blood pressure, for example. Recent scientific evidence is also beginning to show that even some saturated fats, like those in 2% milk, are not all bad. So much for all those years that I spent drinking tasteless skim milk! Anyways, the science and information that we have is always changing, and consequently Canada's food guide also changes to keep up with new scientific evidence to try to keep Canadians healthy and health costs down.

Glycogen Metabolism: Insulin

Insulin is released in response to elevated glucose levels circulating in the bloodstream following a meal. It signals the circulating glucose to be taken up into tissues to be used for energy or to be stored away for later use. When insulin binds to its receptor at the surface of the cell, it signals a series of cellular events that result in the activation of glycogen synthase and the concurrent inactivation of glycogen phosphorylase because the circumstances of the organism dictate that much glucose has become available from a meal. This glucose can be used for energy, therefore, no need for glycogen phosphorylase to be active. At the same time, it signals an abundance of glucose that can be used to replenish the glycogen stores, and therefore signals the activation of glycogen synthase.

ATP is the energy currency of the cell.

It is broken down to ADP and inorganic phosphate (Pi), while releasing free energy that drives certain reactions or processes inside the cell.

PDH Is Also Regulated By Competitive Inhibition

Like glycogen synthase, PDH is also regulated by feedback inhibition when concentrations of acetylCoA and NADH are elevated. These serve to feedback and inhibit PDH when they accumulate in the cell.

Covalent Regulation Of PDH By Phosphorylation

Like glycogen synthase, PDH is regulated through covalent regulation by a similar phosphorylation/dephosphorylation cycle. When it is phosphorylated by PDH kinase, it is inactivated. Conversely, when it is dephosphorylated by PDH phosphatase, it is activated

The Electron Transport Chain

NADH and FADH2 will donate their electrons to the electron transport chain. The electron transport chain is made up of five protein complexes which are embedded within the inner mitochondrial membrane. These are named protein complex I, II, III IV and V (I know...very creative. But at least we don't have to learn a bunch of more complicated scientific names for them!). In general (now please note that I said "in general"...because as we have seen so far, there are often exceptions to some of the fundamental principles in metabolism), the role of the first four protein complexes is two-fold: 1. to accept electrons and shuttle them along, and 2. to pump protons out of the mitochondrial matrix into the intermembrane space. We will discuss these pesky exceptions in the next little while. There are also two electron carriers (coenzyme Q and cytochrome c) which serve to shuttle the electrons between protein complexes. The reason for the carriers is that the electrons do not diffuse very well on their own through the membrane (remember than an electron in its own is charged and thus will not traverse through hydrophobic interior of the membrane), therefore, we need these carriers to shuttle them from one protein complex to the next. Fortunately, coenzyme Q and cytochrome c are lipid soluble and can fulfill this function.

How Are Electrons Shuttled Through The Electron Transport Chain?

NADH donates its electrons to complex I. Then complex I passes these along to complex III then to complex IV. Recall that FADH2 is the coenzyme in the citric acid cycle that is a prosthetic group that is covalently bound to the enzyme succinate dehydrogenase which is actually part of complex II. Wow, small world, eh?! I like to see when all of these pathways begin to come together into this complex and interconnected web. Metabolism is so cool! Hopefully you agree with me by the end of the course! Okay, back to FADH2... FADH2 donates its electrons to complex II (which is both logical and convenient since it is already there anyways...), then complex II passes these along to complex III then to complex IV. Recall that the job of the two carriers is to shuttle electrons through the hydrophobic membrane interior between complexes: coenzyme Q shuttles the electrons through the membrane to complex III and cytochrome c shuttles the electrons from complex III to complex IV. The 'shuttling' of electrons is driven by the reduction potential of the coenzymes and each of the individual redox centres until the final electron acceptor, which is oxygen. This final reduction of oxygen is catalyzed by the enzyme, cytochrome oxidase, which is part of complex IV. The reduction potential can be defined as the affinity that something has for electrons. At the beginning of the electron transport chain, we see that NADH has the lowest reduction potential (or the least affinity for electrons), whereas oxygen has the highest reduction potential (or the highest affinity for electrons). All of the redox centres in between NADH and oxygen have an increasing reduction potential as you pass from protein complex I to III to IV and finally to oxygen. So this is why we need oxygen in the air that we breathe! It is required for the final step in the process of metabolism, and this is why we refer to this process, namely, the electron transport chain and the citric acid cycle that generates the reducing equivalents for the electron transport chain, as 'oxidative phosphorylation'! 'Oxidative' requires oxygen, and the fuels that we use, like glucose and fat, are the compounds that are being 'oxidized'. 4 COMPLEXES: NADH to Ubiquinone, Succinate to Ubiquinone, Ubiquinone to Cytochrome c, Cytrochrome c to O2

Yield Of ATP Per NADH And FADH2

Note that complex I and III each pumps 4 protons, and complex IV pumps 2 protons. Complex II does not pump any protons. So, for each NADH that donates its electrons to the electron transport chain at complex I, a total of 10 protons are pumped out. For each FADH2 that donates its electrons through complex II, a total of only 6 protons are pumped out. As a result, NADH yields 3 ATP, while FADH2 yields only 2 ATP in comparison. These numbers are referred to as the P/O ratios for NADH and FADH2, meaning that for each mole of oxygen that is consumed, three high energy phosphate bonds are generated per NADH and two high energy phosphate bonds are generated per FADH2. Now, you may also have read in different textbooks and sources that a more accurate number of ATP is 2.5 ATP per NADH and 1.5 ATP per FADH2, but for the purpose of this course, we will use nice round numbers that will help make the mental math much easier if we ask you to calculate the number of ATP that can be generated from the oxidation of one molecule of glucose.

Beta Oxidation Of Fatty Acids In Matrix

Now that the fatty acid has been activated and transported into the mitochondrial matrix, it can now be broken down through beta oxidation. The enzymes of beta oxidation are all located in the matrix. One round of beta oxidation involves the sequential removal of two carbons at a time from the acyl group. It is referred to as beta oxidation because it cleaves the carboncarbon bond that is adjacent to the beta carbon in the Greek numbering system. The red dashed line indicates the bond that is being cleaved in one round of beta oxidation. The beta carbon is the third carbon in the acyl group. As you can see, when you cleave the carbon-carbon bond as indicated by the red dashed line, you will generate an acetylCoA molecule! But the process of cleaving that bond is not that simple! It actually involves a series of four reactions that occur in a single round of beta oxidation (you can see Figure 16.15 in the text book if you would like to see the detailed mechanism of beta oxidation - however, for the purpose of this course, you need only know the level of detail that we discuss in this video). One of these reactions will generate an FADH2 and another one will generate NADH. Remember that "beta oxidation" is an oxidative process. It has the word "oxidation" in the name after all!

How Many ATP Are Generated From The Breakdown Of Glucose?

Okay, let's count some beans! What I mean by 'bean counting' is: let's add up how many ATP are generated from the oxidation of one molecule of glucose. As you can see, the partial oxidation of glucose solely through substrate level phosphorylation through glycolysis, which is independent of oxygen (otherwise known as anaerobic metabolism), leads to the production of a net of 2 ATP. As the rest of the original glucose molecule is fully oxidized by the pyruvate dehydrogenase reaction and the citric acid cycle, there is a further 2 GTP (equivalent to 2 ATP) that are generated. But glycolysis also produces 2 NADH. The pyruvate dehydrogenase reaction produces a further 2 NADH, and the citric acid cycle produces 6 more NADH and 2 FADH2. If we multiply each NADH by 3 ATP per NADH, and if we multiply each FADH2 by 2 ATP per FADH2, then we have a total of 38 ATP generated per glucose molecule that is oxidized

Glycolysis: Net Reaction

Once glucose enters the cell from the bloodstream, it can be oxidized to provide useable energy to the cell in the form of ATP. Glycolysis is the sum of 10 different reactions that convert glucose to pyruvate. Note that the 6-carbon glucose molecule is split in two during glycolysis and therefore yields two three-carbon pyruvate molecules, in addition to 2 ATP molecules and 2 NADH molecules. Pyruvate can then further be oxidized in the mitochondria. NADH can also be oxidized by the electron transport chain inside the mitochondria. Other monosaccharides like fructose and galactose also feed into glycolysis to be oxidized

Strategy 1: Formation of Lactate

One form of fermentation that is employed by microorganisms and some eukaryotic cells is the formation of lactate. You are probably very familiar with the concept that intense exercise results in the formation of lactate which can contribute to muscle soreness and fatigue. The enzyme lactate dehydrogenase oxidizes NADH, reducing pyruvate to lactate, and regenerating NAD+ which will allow glycolysis to continue.

Covalent Regulation Of PDH By Phosphorylation continued

PDH kinase is sensitive to regulation by the following effectors: acetylCoA, ATP and NADH are activators of PDH kinase. Pyruvate and ADP are inhibitors of PDH kinase. Insulin and Ca2+ are activators of PDH phosphatase. Elevated concentrations of acetylCoA, ATP and NADH stimulate the PDH kinase and therefore promote phosphorylation and the concomitant inactivation of PDH, whereas, elevated concentrations of pyruvate and ADP inhibit the kinase and therefore promote the active form of PDH, PDHa. Elevated levels of acetylCoA and NADH signal to PDH that they have accumulated in the cell and therefore, there is no need to further breakdown pyruvate until they can be utilized further downstream. Elevated levels of ATP signal that the energy status of the cell is high and that no further breakdown of pyruvate is necessary. Conversely, elevated levels of pyruvate signal a feed-forward mechanism to stimulate PDH to breakdown the pyruvate that has accumulated. Elevated levels of ADP signal that the energy status of the cell is low and that pyruvate should be oxidized for energy. PDH phosphatase is activated by insulin and Ca2+ . Insulin is released following consumption of a meal. This signals that glucose is abundant and can be broken down for energy, and any excess glucose can be converted to acetylCoA which is the precursor for fat synthesis. Through this mechanism, excess glucose will be converted to fat for longer term storage. Ca2+ is released during exercise to signal muscle contraction. By activating PDH phosphatase, it will promote activation of PDH to its active form to increase flux of pyruvate into the citric acid cycle for the continued generation of ATP to support the exercising muscle. It is the relative balance of each of these positive and negative effectors which will determine whether PDH exists in its more active a-form or less active b-form.

Pyruvate Dehydrogenase Enzyme Is A Multienzyme Complex

Pyruvate dehydrogenase is actually a multi-enzyme complex. It is made up of three core enzyme subunits, named E1, E2 and E3, which participate in the oxidative decarboxylation of pyruvate. Lipoic acid is an important coenzyme that is covalently attached to the E2 enzyme that serves as a 'swinging arm' for the acetyl group as it goes from one enzymatic reaction to the next in the overall reaction of the pyruvate dehydrogenase complex. The pyruvate dehydrogenase multi-enzyme complex is also composed of two regulatory enzymes (PDH kinase, which inactivates PDH by phosphorylation, as well as PDH phosphatase, which activates PDH by dephosphorylation)

Structure Of Coenzyme A (CoASH)

Recall that coenzyme A is just that, a coenzyme that participates in the pyruvate dehydrogenase reaction. It is a derivative of pantothenic acid, which is a B vitamin. The free thiol group is the reactive part of coenzyme A which will form an energy-rich thioester bond with the two-carbon acetyl group that will be derived from pyruvate. In this reduced form, coenzyme A is also referred to as CoASH. You need not memorize this structure, but I wanted you to get an idea of how big the coenzyme is, relative to the tiny little two-carbon acetyl group that it will carry!

Triacylglycerols Are The Storage Form Of Fats

Recall that fats are stored in the body in the form of triacylglycerols. The term "triglyceride" is an alternative name for triacylglycerols. Triacylglycerols are made up of three fatty acids which have been esterified with glycerol. Triacylglycerols are usually made up of two or more different types of fatty acids. Most of our fat stores are found in our adipose tissue in specialized cells called "adipocytes". Some tissues like muscle also have a small amount of fat stored within the cell.

Regulation of F6P and F1,6P Cycle

Recall that phosphofructokinase is activated by AMP and fructose-2,6-bisphosphate. Therefore, when the energy state of the cell is high (high levels of ATP and low levels of AMP), gluconeogenesis is stimulated, whereas when the energy state of the cell is low (how levels of ATP and high levels of ADP and AMP), glycolysis is stimulated. Conditions favour the synthesis of either fructose-6-phosphate or fructose-1,2- bisphosphate, but never both at the same time. This fructose-6-phosphate and fructose-1,6-bisphosphate cycle illustrates how glycolysis and gluconeogenesis are reciprocally regulated and do not operate at the same time to avoid futile cycling

Nutrient Breakdown Generates Energy

Recall that there are three macronutrients that provide fuel to the cells for energy production: Carbohydrates (which include polysaccharides like starch that are broken down to glucose), lipids and proteins. Glucose is broken down to pyruvate through glycolysis, then to acetylCoA via the pyruvate dehydrogenase reaction, then to carbon dioxide through the citric acid cycle. The reducing equivalents, NADH and FADH2, that are generated from these processes are then converted to ATP through the electron transport chain and oxidative phosphorylation. Fats are broken down from triacylglycerols to free glycerol and fatty acids. The glycerol will feed into glycolysis, while the fatty acids will be broken down to acetylCoA through beta oxidation. The acetylCoA will then feed into the citric acid cycle, just as acetylCoA derived from pyruvate does. Again, the reducing equivalents that are generated from beta-oxidation (see yellow arrow) and the citric acid cycle are then converted to ATP through the electron transport chain and oxidative phosphorylation. Proteins are broken down to their individual amino acids, and will feed into the above pathways, depending on the amino acid. As you can see, all three paths for macronutrient breakdown converge into an intricate web of linked and integrated common pathways.

Mechanism Of ATP Synthesis

Recall that there is an electrochemical chemiosmotic gradient with a high concentration of protons on the outside of the inner membrane and a relatively lower concentration of protons inside the matrix. This is potential energy that can now be converted to ATP, which is usable energy for the cell. When protons pass through the F0 channel and stalk from the outside to the inside of the matrix, this drives the ATP synthase in the F1 subunit. The proton serves to drive the molecular motor of the ATP synthase which has an ADP and an inorganic phosphate (Pi) present in the active site. The proton actually drives the conformational change in the active site, and hence drives the catalysis of the formation of a new high energy phosphoanhydride bond in ATP.

Shuttling Electrons Through The Electron Transport Chain

So exactly what is it inside these protein complexes that accepts the electrons? It is not the proteins in the complexes that are reversibly reduced and re-oxidized. But, within each protein complex there are several redox centres which can be reversibly reduced and re-oxidized, just like NADH and FADH2 can be reoxidized to NAD+ and FAD when they donate their electrons. These redox centres can be primarily categorized as: 1. Coenzymes, 2. Fe-S clusters, 3. cytochromes and 4. Cu. Each protein complex contains a combination of two or more of these types of redox centres.

Muscle Metabolism From Rest To Exercise

So let's use muscle as a model to study and understand metabolism since its demands can vary tremendously from rest to high intensity short-term sprint exercise or low intensity longer-term marathon running. Recall that the preferred fuel for muscle at rest is fat. Production of ATP from fat is able to meet the demand for ATP at rest and at very low exercise intensities, such as a leisurely stroll through the park. The fats are delivered to the muscle via the bloodstream from the adipose tissue as a constant supply, just like the heart. As we increase our exercise intensity, the demand for ATP increases and the rate of production of ATP from the oxidation of fats is no longer able to meet the demand as the pathway of beta oxidation and the citric acid cycle are slower processes. Therefore the body begins to rely more heavily on the oxidation of glucose. Glycolysis provides ATP through substrate level phosphorylation at a much greater rate than the flux of metabolic intermediates through beta oxidation or the citric acid cycle and oxidative phosphorylation. At intermediate exercise intensities, the body relies on a blend of fat and carbohydrate degradation. The fatty acids continue to be supplied via the bloodstream, while the glucose is also being provided through the bloodstream, originating from the liver. As we increase our exercise intensity even further to a sprint, fat oxidation provides very little ATP, whereas the breakdown of glucose through glycolysis provides the greatest proportion of ATP along with a high energy phosphate compound called creatine phosphate. Now, the supply of glucose from the bloodstream (originating from the liver glycogen stores) is not able to keep up with the demand, therefore, the muscle turns to its own local and immediate stores of glycogen for a rapid supply of glucose for glycolysis. Now how about that creatine phosphate that I just mentioned?

Overview Of Fat Metabolism

So now that we have gotten over our aversion to fats, let's look at how it is metabolised in the body! Remember that there are anabolic processes and catabolic processes. In carbohydrate metabolism we compared and contrasted anabolic vs catabolic process like glycogen synthesis vs glycogen breakdown, and gluconeogenesis vs glycolysis. The same holds true with fat metabolism. We will look at both fat synthesis and fat breakdown. This schematic shows the interplay and interactions of some common metabolic intermediates that we have already seen, such as pyruvate and acetylCoA, which were derived from glucose breakdown. In this module, we will see that there are other sources of acetylCoA, such as from the breakdown of fats (triacylglycerols), ketone bodies and some amino acids. Through common intermediates like pyruvate and acetylCoA, multiple pathways are interconnected. Let's first take a look at the breakdown of fats

Fatty Acid Biosynthesis

So that was the catabolic process of fat breakdown. Now let's switch gears and look at the anabolic process of fat synthesis. In this case, unlike gluconeogenesis and glycolysis which share some of the same enzymes in each pathway, fat synthesis does not share any of the same enzymes that take part in beta oxidation. However, conceptually, the process is essentially the reverse of beta oxidation.

Pumping Protons

So the electron transport chain is not just about shuttling electrons. Recall that it is also about pumping protons. Each time electrons pass through a protein complex (with one exception) from a redox center with a lower reduction potential to a redox center with a higher reduction potential, energy is released. The protein complex harnesses that energy and uses it to drive a proton pumping mechanism that pumps protons out of the matrix and into the intermembrane space. Complex I and complex III each pumps out four protons for each pair of electrons that pass through, while complex IV pumps only two protons. Complex II does not actually pump protons.

Lactate Production During Exercise

So the take home message is that, as exercise intensity increases, we shift from a reliance on fat oxidation to a greater reliance on glycolysis of glucose. The downside of increasing reliance on ATP production from substrate level phosphorylation from glycolysis is that lactate is generated as a bi-product of metabolism when the NAD+ must be regenerated in order to allow glycolysis to continue during high intensity exercise. Lactate results in muscle fatigue and cramping and reduces our ability to continue to exercise at a high level of intensity. So this is why we can't keep up a sprint for a long period of time. Eventually, the lactate produced slows us down and we exhaust our creatine phosphate stores as well.

Regulation Of Glucose Metabolism

So what are the regulatory signals that keep a constant supply of glucose in the bloodstream in between meals, and takes up excess glucose after a meal? This is very important as we need to keep a constant supply of glucose for our all-important brain! Post-prandially (that is, after a meal), there are elevated levels of glucose in the circulation. That stimulates the release of insulin from the pancreas. This signals the uptake of glucose into the tissues, namely muscle and liver. The glucose is stored away as glycogen. (Recall that any excess glucose is converted to fat and stored for longer term use.) A few hours post-prandially (after eating), our blood glucose levels begin to drop a little bit. In order to maintain glucose homeostasis in the blood, the pancreas releases the counter regulatory hormone to insulin, which is glucagon. Glucagon stimulates glycogen breakdown in the liver to maintain a constant supply of glucose in the bloodstream. When there is a dysfunction in insulin signaling, this leads to type two diabetes. It is a bit of a puzzle and may be multifactorial, and scientists are struggling to understand and treat it as it bears a significant burden on the health care system with increasing incidence in recent decades. Because muscle occupies approximately half of our muscle mass, it is a significant consumer of glucose, and regular exercise can help maintain healthy glucose levels and prevent the development of type two diabetes.

starvation

So what happens after you have been fasting for a much longer period of time. We are talking longer than in between meals - days and weeks... Any longer than about half a day or so, and our liver glycogen stores have likely been pretty much depleted. Now what? Well, the brain isn't going to be happy, but fortunately our body usually has fat stores in reserve in our adipose tissue (as I have mentioned, some of us have more fat stores than others...). Those fatty acids can be taken up from the bloodstream by the liver and be converted to ketone bodies. While the body isn't exactly excited about ketone bodies, it is better than nothing. Under periods of starvation, the brain (and heart too) will adapt to using ketone bodies. Now, our fat stores are quite significant in terms of energy production, but they are limited. So what happens when we have exhausted all of our adipose tissue reserves? The last and not preferred avenue is protein. While our body does not store protein, per se, in the same way that it stores glucose and fat, we do have significant protein reserves in our muscle mass. Protein can be broken down to its amino acid components, and the glucogenic amino acids will be converted to glucose and the ketogenic amino acids will be converted to ketone bodies. So why not break down muscle protein since some of those amino acids are glucogenic and can generate the glucose that the brain prefers? Our bodies will preserve our muscle mass until it has no other options. Because if we broke down muscle protein as a first option, before breaking down fats and synthesizing ketone bodies, we would not be able to move around to gather food. It is a mechanism of self-preservation and survival. But when there is no fat left, the body has no other option but to break down muscle proteins to generate ATP for our cells to survive.

Fuel Metabolism: Preferred Fuels

So what is the preferred fuel for each tissue or organ in our body? Now these are just the 'preferred fuels'. As we will see in the upcoming graphics and images, some tissues and organs can make use of other fuels if needed. Recall that the brain prefers glucose (but during starvation, it will gladly accept ketone bodies...). At rest, the muscle is very content to burn fatty acids, but during exercise, will switch over to burning more glucose. The heart is always pumping, and therefore is never really "at rest" in the same way that skeletal muscle is, and therefore prefers to get its fuel from the constant and steady flow of fatty acids which are a rich source of energy. Adipose tissue, as we may expect, burns fatty acids since that is what its specialty is! Lastly, the liver is the 'jack of all trades' in the world of metabolism and is not that picky and will burn any of the three macronutrients to produce energy.

Fuel Metabolism: Fuel Reserves

So where do we store the majority of our fuels in our body? The brain doesn't actually keep anything on reserve. It is the liver's job to store glucose as glycogen for the entire body to use over the course of the day. Which is why breakfast is the most important meal of the day. Dinner the night before was a long time ago, and the liver's reserve of glycogen is running low. You need that brainpower to perform well at school! One of the liver's main functions is to process all of the fats that are ingested after a meal. It repackages it as chylomicrons and sends it out into the circulation for other tissues to pick up as a fuel. It has a little bit of triacylglycerols stored within, but not too much as you don't want a fatty liver. That is a bad thing. Most of our fats are stored in our adipose tissue for longer term storage. And that is why bears want to pack away some extra fat reserves before a long winter of hibernation, and why you should also pack away some extra fat reserves if you want to be a contestant on the television show Survivor if you hope to survive 39 days on eating berries and cockroach delicacies. Ick! The heart doesn't actually have any energy reserves of its own. It relies on the circulating fatty acids in our blood stream to provide it with a constant supply of fuel. Skeletal muscle on the other hand occupies approximately half of our total body mass and also has its own carbohydrate reserves as glycogen and a very small amount of triacylglycerols as well. Let's talk about muscle metabolism as an illustration of how our body draws on these different fuel reserves.

Activation Of Fatty Acids In Cytosol

So why do we need to "activate" the fatty acid? Recall the reaction of pyruvate dehydrogenase that converted the pyruvate to acetylCoA... The role of the coenzyme A was to "activate" or raise the energy of the acetyl group by forming a high energy thioester bond and preparing it to enter into the citric acid cycle for oxidation. Well, this is exactly what we are doing here. In this case, rather than attaching a short two-carbon acetyl group to the coenzyme A, we are attaching a long chain hydrocarbon of a fatty acid to coenzyme A. Same idea... The enzyme that catalyzes this reaction is an acylCoA synthetase. The "acyl group" is the portion of the fatty acid that is bound to the coenzyme A. This reaction takes place in the cytosol, and the acylCoA is now ready and primed to be transported into the mitochondrial matrix to be oxidized.

Some Common Themes In Metabolic Regulation

So, a common theme in the regulation of the citric acid cycle is very similar to what we have seen in the regulation of the glycolytic pathway: that the energy status of the cell, reflected by the relative concentrations of ADP and ATP, and the redox state of the cell, reflected by the relative concentrations of NADH and NAD+ are important regulators of metabolism. In addition, it is common for regulatory enzymes upstream in a pathway to be subject to feedback inhibition by products. This serves to fine-tune flux through the pathway overall. Again, not every enzyme in a pathway is regulated, but there are normally two or three key enzymes in a pathway that represent points of regulation.

Fatty Acid Metabolism: High Glucose

So, how is fat metabolism regulated? Under conditions when glucose is high, for example, after a meal rich in carbohydrate, insulin is released. The hormone insulin activates fat synthesis in the liver and simultaneously inhibits fat breakdown in adipose tissue. This is a good thing. Remember the concept of futile cycling? We want to avoid breaking down fats when glucose is plentiful and the body is in the mode of synthesizing fats. Remember that it is energetically expensive to synthesize fats. It costs 7 ATP to produce one molecule of palmitate!!

Control of Flux Through Glycolysis step 1

Step 1 is catalyzed by the enzyme hexokinase. It is inhibited by its product, glucose-6- phosphate, via end-product inhibition. High concentrations of glucose-6-phosphate signal that there is enough substrate for glycolysis to proceed, and that additional glucose need not be broken down at the moment.

Control of Flux Through Glycolysis step 3

Step 3 is catalyzed by the enzyme phosphofructokinase. It is inhibited by high concentrations of ATP and citrate. This is another form of end-product inhibition whereby the end products of a pathway feedback and inhibit flux through an enzyme. The role of ATP as an inhibitor is logical as it signals that the cell has adequate levels of ATP and that flux through glycolysis can be down-regulated. Citrate is the first product in the tricarboxylic acid cycle further downstream from glycolysis. It also signals that there is enough substrate for the tricarboxylic acid cycle to proceed, and therefore further glucose need not be broken down. Conversely, phosphofructokinase is stimulated by high concentrations of AMP and ADP which signal a reduction in the energy status of the cell, meaning that the concentration of ATP is low. Its activity is also up-regulated allosterically by an important metabolite, fructose-2,6-bisphosphate.

Ketone Bodies: Brain Fuel During Fasting

Thankfully, the ketone bodies produced in the liver during fasting and starvation enter into the bloodstream and are able to cross the blood:brain barrier and be taken up by the brain and broken back down into acetylCoA, which can then enter the citric acid cycle and produce energy

In the case of epinephrine binding to the beta adrenergic receptor, the transducer is a G protein.

The G protein is a heterotrimeric protein (that is, a protein made up of three different subunits - the alpha, beta and gamma subunits). It reversibly binds GDP and GTP. Binding of the hormone to its receptor induces a conformational change in the receptor which promotes interaction with the G protein (see left panel) and stimulates exchange of the bound GDP for GTP (see right panel for details). When GDP is bound to the G protein, it is in its inactive state. When GTP is bound to the G protein, it is in its active dissociated state. The active form of the G protein (rendered active by reception of the hormonal signal) will then subsequently activate the effector, an enzyme called adenylate cyclase, to elicit the cellular response

Electrons From Cytosolic NADH Must Pass Through A Shuttle

The NADH that is generated inside the mitochondria through the pyruvate dehydrogenase reaction and the citric acid cycle will have easy access to the electron transport chain to donate its electrons. However, the NADH that are generated from glycolysis which takes place in the cytosol will not have access to the electron transport chain to donate its electrons. Recall that the inner membrane is relatively impermeable, and therefore larger and charged molecules must be transported across the membrane via specific transport proteins. While there is no actual transporter for NADH (NADH never actually crosses the inner membrane), there are two mechanisms by which NADH can donate its electrons to another intermediate which can then traverse the membrane. While you don't need to know the details of these two mechanisms, you just need to be aware that the glycerol3-phosphate shuttle and the malate-aspartate shuttle are the two pathways that provide the electrons from NADH generated in the cytosol to the electron transport chain.

Cell response: part 1

The active G protein interacts with and activates the "effector" which is an enzyme called adenylate cyclase (AC) (see left panel). Adenylate cyclase catalyzes the conversion of ATP to cAMP (see right panel). cAMP is referred to as a second messenger. In this example of signal transduction, the hormone, epinephrine, was the first messenger. So before we move on, let's take a step back and put everything together. The hormone, epinephrine, binds to its beta adrenergic receptor. It interacts with and activates the G protein, which then interacts with and activates adenylate cyclase. Make sure that you understand the three steps of signal transduction and make sure that you can identify the signal, transducer, effector and second messenger in this scenario.

Advantages Of Multienzyme Complexes

The advantages of multienzyme complexes such as pyruvate dehydrogenase are as follows: 1.There is an increased rate or efficiency of reaction due to a reduction in the diffusion distance for intermediates between consecutive enzymes 2.The complex channels intermediates between successive enzymes in a pathway, minimizing side reactions (meaning that the reaction goes to completion). In fact, this is the function of the lipoic acid 'swinging arm mechanism' - to channel the substrate from enzyme to enzyme. 3.Lastly, reactions catalyzed by multienzyme complexes can be coordinately regulated

Gluconeogenesis is Energetically Favourable

The answer is simple. Gluconeogenesis is not exactly the reverse of glycolysis. Those three non-equilibrium reactions in glycolysis which have large changes in free energy are by-passed in gluconeogenesis. The other reactions are near-equilibrium reactions and therefore are shared between the two pathways. After all, sharing is caring, and that makes good sense in a cell that is packed full of enzymes and reactions. As a result, energy input is required for gluconeogenesis to proceed. This makes good sense since glycolysis generates energy, then we would expect that the reverse would require energy input. Also, the concentrations of substrates are different under the metabolic circumstances that occur when gluconeogenesis is required, thus driving the near-equilibrium reactions in the reverse direction

Signalling pathways involving the insulin receptor

The beta adrenergic receptor that we have just reviewed belongs to a class of G-protein coupled receptors (or GPCRs for short). There are many different types of receptors involved in signal transduction, so let's briefly look at one more type: the receptor tyrosine kinase family. These receptors are so named because they each have a protein tyrosine kinase domain which is activated when the ligand binds. One such receptor is the insulin receptor. When insulin binds to its receptor, it activates the tyrosine kinase activity which then activates another cascade mechanism whereby PIP3, another second messenger, is synthesized. Eventually at the end of the cascade, GLUT4 (or glucose transporter 4) proteins are relocalized to the plasma membrane and are responsible for taking up glucose from the blood stream after a meal. So, in this case, insulin is the signal, or messenger, the insulin receptor receives the signal, it is then transduced to the inside of the cell to effect the cellular response which is the relocalization of GLUT 4 and subsequent glucose uptake.

Cell response: part 2

The cAMP then binds to the cAMP-dependent protein kinase A and subsequently activates it. The active protein kinase A then phosphorylates phosphorylase kinase b. Phosphorylation of phosphorylase kinase b causes it to become active. The phosphorylation/dephosphorylation cycle is common in the activation or deactivation of many enzymes when we talk about the regulation of enzyme activity. The active phosphorylase b kinase then activates glycogen phosphorylase (or phosphorylase for short) by phosphorylating it. In the case of glycogen phosphorylase, it actually has two different names when it is in its inactive vs its active form. We refer to glycogen phosphorylase b as the dephosphorylated inactive form, and we refer to glycogen phosphorylase a as the phosphorylated active form. The active glycogen phosphorylase a then catalyzes the breakdown of glycogen, releasing glucose-1-phosphate for degradation and ATP synthesis to support muscle contraction, which is the reason why epinephrine was released in the first place, when exercise and muscle contraction are initiated

The Citric Acid Cycle: The Reactions -- Movie Narrative

The citric acid cycle is a series of chemical reactions that takes place in the mitochondrial matrix. This cycle uses acetyl CoA, derived from sugar and fat breakdown, to form ATP, NADH, FADH2, and carbon dioxide. The NADH and FADH2 can be used to form additional ATP through the electron transport chain. The citric acid cycle goes by many names including the tricarboxylic acid (or TCA) cycle and the Krebs cycle. Citric acid refers to the citrate that is produced in the first step of the pathway. The tricarboxylic acid title gets its name from the three carbon dioxides that are produced for each fully oxidized pyruvate. Krebs refers to Hans Adolf Krebs who identified the full cycle in 1937. He was eventually awarded the Nobel Prize for Physiology or Medicine in 1953 for his discovery. In the first step of the cycle, an enzyme called citrate synthase joins the two-carbon acetyl group from acetyl CoA with the four-carbon oxaloacetate to form a six-carbon citrate. In step two, an enzyme called aconitase converts citrate into isocitrate. Next, an isocitrate dehydrogenase enzyme oxidizes isocitrate, a six-carbon molecule, to a fivecarbon α-ketoglutarate. The carbon that was lost is released as carbon dioxide and one NADH is also formed. The carbon dioxide that is released was originally part of oxaloacetate and not acetyl CoA. In the fouth step, an enzyme called α-ketoglutarate dehydrogenase converts αketoglutarate into a four-carbon succinyl CoA. Similar to step three, this reaction produces one carbon dioxide and one NADH. In step five, a succinyl CoA synthetase enzyme converts succinyl CoA into succinate. This produces GTP which is converted to ATP. In step six, an enzyme called succinate dehydrogenase converts succinate into fumarate. This step makes one FADH2. In step seven, a fumarate hydratase enzyme then converts fumarate into malate. In the final step of the citric acid cycle, a malate dehydrogenase enzyme converts malate back to oxaloacetate. Like all steps involving a dehydrogenase, a coenzyme is produced. Here it is NADH. The oxaloacetate that was regenerated through the citric acid cycle is now ready to join with another acetyl group and begin the cycle a second time. For every one glucose that is broken down through glycolysis, two pyruvates will be produced. These two pyruvates will produce two acetyl CoAs. So, for every one glucose, two acetyl CoAs will be made and two turns of the citric acid cycle will occur. This means each product of the cycle must be doubled. A total of four CO2, six NADHs, two FADH2, and two ATPs are made through the citric acid cycle. NADH and FADH2 are electron carriers that can produce more ATPs later in aerobic respiration. In addition to sugars like glucose, proteins and fats can also provide carbon substrates to fuel the citric acid cycle. Proteins can be broken down into individual amino acids such as alanine, aspartate, and glutamate and converted into intermediates in the cycle. Fatty acids can be broken down into acetyl CoA which then begins the citric acid cycle. This metabolism of sugars, proteins, and fats through the citric acid cycle provides the vital energy necessary to maintain many cellular processes

names of citric acid cycle

The citric acid cycle is also know as the tricarboxylic acid cycle and the Krebs cycle. So this is one of the many fun things that you sometimes find in science - when there is more than one name for the same thing! The origin of the name is of course attributed to the scientist who discovered the cycle in 1937. It is also named after the product of the first reaction, which is citrate. And citrate is a tricarboxylic acid! (But you need not memorize this structure.) And the easy part is the last word, 'cycle', because it begins and ends with oxaloacetate

Regulation Of The Citric Acid Cycle

The citric acid cycle is subject to regulation at many levels. Again, not all enzymes in this cycle are regulatory in nature. Just a few enzymes in a pathway need to be regulated in order to control flux. There are three ways in which the key regulatory enzymes in the citric acid cycle are regulated: 1.Availability of substrates 2.Competitive inhibition by products 3.Allosteric regulation First, the availability of both initial substrates is important. Oxaloacetate and acetylCoA must be available in a 1:1 ratio for this first reaction to occur. There must also be an adequate supply of free NAD+ to allow the citric acid cycle to continue, much like free NAD+ is required for glycolysis to continue. Second, an accumulation of products, such as NADH, will inhibit some of the key enzymes in the pathway, such as the dehydrogenases that are responsible for generating these reducing equivalents. Lastly, some key enzymes are subject to positive and/or negative allosteric regulation by allosteric effectors such as calcium ions. Looking at skeletal muscle as an example, the release of calcium ions stimulates muscle contraction, while at the same time signalling the breakdown of fuel for energy production by stimulating some key regulatory enzymes in the citric acid cycle. It is a very clever coupling of muscle contraction with energy generation to meet the energy demands during exercise! Our cells are so smart!! Calcium is an example of positive allosteric regulation in the citric acid cycle. ATP is an example of a negative allosteric regulator which signals that the energy status of the cell is high, and that the citric acid cycle need not oxidize any further acetylCoA to meet the energy needs of the cell. Conversely, we see that ADP is a positive allosteric regulator as it signals that the energy status of the cell is low, and ATP production needs to be ramped up through increased flux through the citric acid cycle.

Creation Of A Chemiosmotic Gradient

The effect of shuttling electrons, combined with proton pumping, has now created a chemiosmotic gradient across the inner mitochondrial membrane. Due to donation of electrons from NADH or FADH2 to the electron transport chain, there is high concentration of protons outside of the inner membrane, relative to the inside of the matrix. Recall that the inner mitochondrial membrane is relatively impermeable. It is impermeable to compounds that are charged and/or large. Therefore, the only way for positively charged protons to traverse the inner mitochondrial membrane is through a specific transport protein. The nature of the gradient is referred to as 'electrochemical' because it is both chemical in nature in terms of creating a lower pH outside of the inner membrane, and 'electrical' in nature because the positive charge of the protons wish to reach equilibrium by entering the mitochondrial matrix where the relative charge is negative. Now, keep in mind that these are localized gradients along the inner membrane on the outside and inside of the matrix where proton pumping occurs. There is potential energy stored in this chemiosmotic gradient. The cell harnesses that potential energy and uses it to drive the synthesis of ATP by a process referred to as 'oxidative phosphorylation of ADP to ATP' within complex V.

Synthesis Of Fatty Acids In Cytosol

The enzyme called fatty acid synthase is responsible for catalyzing the synthesis of fatty acids. It will take 8 acetylCoA molecules and condense them in a similar rinse and repeat cycle as with beta oxidation, but in reverse, and synthesize the 16-carbon fatty acid called palmitate.

Fates Of The Products Of Triacylglycerol Breakdown

The fatty acids that are generated from the action of lipase leave the adipose tissue and enter the bloodstream to be delivered to the tissues for the purpose of generating energy. Fatty acids are carried through the bloodstream bound to the protein albumin, which keeps the fatty acids soluble. The fatty acids will be taken up by liver and muscle, for example, but not by the brain. Recall that the preferred fuel for the brain is glucose (And is the reason that breakfast is the most important meal of the day! Students can't survive on caffeine alone!)! And under conditions of starvation, the brain will adapt to using ketone bodies. The fatty acids that are taken up into tissues are broken down to acetylCoA via the beta oxidation pathway. The glycerol moiety is also usable energy, and leaves the adipose tissue to be taken up by the liver, which is where gluconeogenesis takes place. Glycerol is a substrate for gluconeogenesis, entering the gluconeogenic pathway at a midpoint

Glycogen phosphorylase is activated by high concentrations of AMP.

The high concentration of AMP indicates that the energy status of the cell is quite low (meaning that the concentration of ATP is reciprocally low), and therefore, signals an increase in the activity of glycogen phosphorylase to release more glucose residues in order for glycolysis to produce more ATP. Conversely, glycogen phosphorylase is inhibited by high concentrations of ATP. The high concentration of ATP signals to the cell that there is ample energy supply to meet the demand, and no further substrate needs to be broken down and utilized.

Glycogen Metabolism: Glucagon

The hormone glucagon, is counter-regulatory to insulin, meaning that glucagon is released when glucose levels in the bloodstream drop, and signals more glucose be released from the liver glycogen stores to restore blood glucose levels. As a result, glucagon inactivates glycogen synthase and activates glycogen phosphorylase

the brain prefers glucose over any other fuel

The muscle also loves glucose during high intensity exercise, but at rest or during low intensity exercise, it is satisfied by the breakdown of lipids.

Pentose Phophate Pathway Makes NADPH

The pentose phosphate pathway is an alternate pathway by which glucose is broken down to generate NADPH for reductive biosynthetic processes such as fat synthesis, as well as providing ribose-5-phopshate for the biosynthesis of nucleotides, depending on the needs of the cell. Here we see the oxidative phase where glucose-6-phosphate is oxidized, generating NADPH and the waste product CO2.

Anatomy Of The Mitochondrion

The pyruvate dehydrogenase reaction and the citric acid cycle are compartmentalized inside the mitochondrion. The mitochondrion is an organelle that specializes in energy production. It is considered to be the "powerhouse of the cell" because much of the ATP that is generated in the cell originates from the mitochondrion. The mitochondrion is encapsulated by a double membrane. The outer mitochondrial membrane surrounds the entire structure, while the inner mitochondrial membrane is invaginated to increase the surface area of the membrane. The invaginations are called "cristae". The inner compartment is called the "matrix". The space between the inner and outer membrane is called the "intermembrane space". The outer mitochondrial membrane is highly permeable with large channels that span the membrane that are called "porins". Porins allow compounds less than approximately 5000 daltons in size to pass. The inner mitochondrial membrane is relatively impermeable, allowing only small uncharged compounds like CO2 and water to cross the membrane. Any larger or charged molecules, such as pyruvate, protons or ATP, will cross the membrane through the presence of specific transport proteins. Compared with the outer membrane, the inner membrane contains a much higher concentration of proteins as it contains these specific transport proteins as well as the protein complexes involved in the electron transport chain. It is the relative permeability of the inner mitochondrial membrane that will allow a proton gradient to be established for the electron transport chain. The pyruvate dehydrogenase enzyme and most (not all) of the enzymes for the citric acid cycle are localized inside the mitochondrial matrix. Hint: we will see that one of the enzymes in the citric acid cycle is part of protein complex II in the electron transport chain which is embedded in the inner mitochondrial membrane.

Cells Require Energy

The reason that we consume food, is to provide fuel to our body to synthesize energy in the form of ATP (but I choose to consume chocolate just because it makes me feel good - there are other terrific chemicals in food like phytochemicals that also lift our mood and make us feel good, but we'll leave that for another course!). Our cells need energy for many things, including housekeeping activities such as DNA replication, cell division, protein synthesis, maintaining an osmotic gradient of sodium and potassium ions... All of these processes require energy.

Signal transduction from start to finish

The signal was epinephrine. The signal was received when it bound to the beta adrenergic receptor (Step 1: reception). The signal was transduced by the G protein (Step 2: transduction). And the effector adenylate cyclase elicited the cell response by synthesizing cAMP which initiated the whole cascade mechanism which led to the eventual activation of glycogen phosphorylase and subsequent breakdown of glycogen (Step 3: cell response).

Coordinate Regulation Of Pyruvate Dehydrogenase And Pyruvate Caboxylase

There is a clever mechanism which coordinates the regulation of pyruvate dehydrogenase and pyruvate carboxylase to ensure a 1:1 ratio of oxolacetate:acetylCoA in the first step of the citric acid cycle. If the supply of acetylCoA (from glycolysis or β-oxidation) is greater than the supply of oxaloacetate, then citrate synthase will not function maximally due to substrate limitation Therefore, excess acetylCoA will inhibit PDH, diverting pyruvate to the pyruvate carboxylase reaction

there is also an opposite pathway whereby pyruvate is converted to glucose.

This is a biosynthetic pathway referred to as an "anabolic" process, or anabolism. Therefore, glycolysis is a catabolic process and gluconeogenesis is an anabolic process.

Fatty Acid Biosynthesis net reaction

This is the net reaction for the synthesis of palmitate. But this is not the only fatty acid that mammals can synthesize. There are also enzymes called elongases and desaturases that make longer chain fatty acids and create double bonds in unsaturated fatty acids. Once the fatty acids are synthesized, they are esterified with glycerol to form triacylglycerols or are used to make membrane lipids.

Cholesterol Synthesis

This is the overall pathway for cholesterol synthesis, just to illustrate how acetylCoA is the 2-carbon donor for the synthesis of cholesterol. The enzyme HMG-CoA reductase is the target for statin drugs that help to lower cholesterol levels in those who are afflicted with high levels of the "bad cholesterol", LDL.

Stage 1: The Pyruvate Dehydrogenase (PDH) Reaction

This is the reaction catalyzed by pyruvate dehydrogenase. It catalyzes the irreversible oxidative decarboxylation of pyruvate to acetylCoA. This process is 'oxidative', because, as you can see, electrons are drawn away, reducing NAD+ to NADH. This is also a 'decarboxylation' reaction because a one-carbon carbon dioxide molecule is removed from the three carbon pyruvate. The carboxyl group of pyruvate is a good 'leaving group' which leaves in the form of carbon dioxide. As you can see here, the free thiol group from CoASH is forming that energy-rich thioester bond (indicated in red) with the two-carbon acetyl group derived from pyruvate. Coenzyme A functions as a carrier which serves to 'activate' the two-carbon acetyl group through the formation of that high-energy thioester bond

Oxidation Of Pyruvate

To recap, the oxidation of pyruvate to carbon dioxide occurs in two stages: First, the decarboxylation of pyruvate to form acetyl-coenzyme A (abbreviated as 'acetylCoA'), followed by the oxidation of the acetyl group of acetylCoA to form carbon dioxide.

Fatty Acid Metabolism: Low glucose

Under conditions when glucose levels are low, such as during fasting or just in between meals, glucagon is released. Glucagon is the counter regulatory hormone to insulin. Its job is to stimulate fat breakdown by stimulating the lipase in adipose tissue. Then fats are broken down to acetylCoA and used for energy, or during periods of extended fasting or starvation, the acetylCoA can be diverted to ketone body synthesis to preserve our very important brain function by providing an alternative form of fuel for the brain. While ketone bodies are not ideal, it is better than nothing at all...

Fermentations: Net Reactions

Under conditions where O2 is absent, fermentation can still continue to produce 2 molecules of ATP to meet the cell's demand for ATP. strategy 1: makes 2 lactate + 2 ATP strategy 2: 2 ethanol + 2 C02 + 2 ATP

Ribose-5-Phosphate is a PPP Intermediate

Under conditions where ribose-5-phosphate is required for nucleotide synthesis, the pentose phosphate pathway will divert carbons to its production.

Electron Transport And ATP Synthesis: The Final Frontier

We are now entering the final stages of energy metabolism: the final frontier (cue Star Trek music...)... We have completely oxidized one molecule of glucose through glycolysis, the pyruvate dehydrogenase reaction and the citric acid cycle. Throughout this catabolic process, we have generated a net of two ATP and two NADH from glycolysis, two NADH from the pyruvate dehydrogenase reaction, and three NADH, one FADH2 and two GTP (remember that these GTP are equivalent to ATP) from the citric acid cycle. The ATP and GTP are usable forms of energy, but there is still lots of energy packed away in those electrons that were passed on to NADH and FADH2 along the way. So now let's take the electrons from those reducing equivalents and convert them to usable energy

Muscle Contraction Is A Major Consumer Of Energy

When we exercise, muscle contraction demands a significant increase in ATP production over and above the day-to-day housekeeping activities of the cell. In fact, as an athlete goes from rest to a full-out sprint, the demand for ATP can increase on average up to 100-fold.

Strategy 2: Formation of Ethanol

You are likely also familiar with the fermentative processes that occur in yeast to make wine and beer. In yeast, ethanol is produced rather than lactate during fermentation which regenerates NAD+ and allows glycolysis to continue.

The three macronutrients that we consume in our diet to generate energy are

carbohydrates (also known as sugars), lipids (also known as fats) and proteins. The three macronutrients that we consume in our diet to generate energy are also stored in the body as energy reserves, with the exception of protein

A pathway is

comprised of a sequential series of reactions in a cell that are catalyzed by enzymes. In the pathway of glycolysis, there are ten different reactions. In this generic Published by Articulate® Storyline www.articulate.com representation of a pathway, the upper case letters denote the molecules in the pathway. These are also referred to as "metabolic intermediates". The upper case letter E with a number over each arrow denotes the enzyme that catalyzes the conversion of one intermediate to the next.

signal transduction

example insulin: The message being communicated is in response to the intake of a meal. In response to the increase in blood sugar levels, the pancreas releases the hormone, insulin, into the bloodstream. Insulin will signal to target cells that they should take up the excess glucose that is circulating through the bloodstream. In this manner, signal transduction communicates a message from the distant pancreas to the target cells in muscle and adipose tissue to take up the glucose and either use it or store it away for future use. The signal molecule, or messenger, (in this case, the insulin hormone is the signal molecule) is transmitted from the pancreas to the target tissues and they will respond accordingly

Glucose is broken down sequentially through

glycolysis, then through the pyruvate dehydrogenase reaction and citric acid cycle .It is important to note that these pathways are highly regulated to control the breakdown of carbohydrates when the body needs it. Glycolysis can proceed in the absence of oxygen, that is, under anaerobic conditions. In humans, this results in lactate production, and in yeast, ethanol is generated as a biproduct of fermentation rather than lactate. Because the brain's preferred fuel is glucose, the body also has the ability to synthesize glucose de novo. This process is called gluconeogenesis. Carbohydrates that are consumed in our diets that are not required immediately for energy are stored as glycogen in our muscle and liver. Once our glycogen stores are filled up, excess carbohydrates are converted to fat for storage. Once again, glycogen degradation and glycogen synthesis are highly regulated processes. Finally, the pentose phosphate pathway is an alternate pathway by which glucose is broken down to generate NADPH for reductive biosynthetic processes such as fat synthesis, as well as providing ribose-5-phopshate for the biosynthesis of nucleotides

classes of extracellular messengers/signals

hormones (act at a distance) neurotransmitters (secretion close to target cells) pheromones (act upon cells in a different organism) growth factors (act at various distances) All of these signal molecules are secreted in small amounts but exert a relatively larger and significant effect through the process of signal transduction. This is referred to as signal amplification and we will look at this mechanism later

The basic building block of protein

is the amino acid unit. As mentioned, the body does not 'store' protein in a way that is analogous to lipid and carbohydrate storage. Nonetheless, under periods of prolonged starvation, the body will draw on the protein found in muscle as energy.

The basic building block of lipids

is the free fatty acid unit. Three of these are esterified with a molecule of glycerol to form a triglyceride, which is how the body stores lipids

The basic building block of carbohydrates

is the glucose unit. Glucose molecules are linked together to form a very large molecule of glycogen, which is how the body stores carbohydrates.

And in biological systems, that energy is largely in the form

of ATP

During catabolic processes

our body breaks down macronutrients through an oxidative process whereby electrons are drawn away and donated to reducing equivalents which are coenzymes like NAD+ , NADP+ and FAD. Enzymes called dehydrogenases catalyze these reactions. We will later see how these reduced coenzymes donate their electrons to the electron transport chain to convert ADP to ATP

We get it from "fuel", and that "fuel" comes from

our diet or fuel reserves in our body

types of hormones

peptides such as insulin or glucagon, steroids such as Vitamin D or estrogen, amino acid derivatives such as epinephrine or thyroxine

Recall that hormones are signal molecules. They act on their target cells by binding to

receptors (often on the surface of the cell, like the insulin receptor, but also sometimes within the cell, like the nuclear receptor for vitamin D).

example of glucagon

secreted by the pancreas and acts on the liver to promote glycogen breakdown and gluconeogenesis when blood glucose levels are low signal molecule

example of insulin

secreted by the pancreas and promotes uptake of glucose into cells when blood glucose levels are high signal molecule

example of epinephrine

secreted from the adrenal gland and promotes the generation of energy in the form of ATP in muscle when it is required for exercise signal molecule

Hormones

small molecules secreted by endocrine glands which travel long distances through the bloodstream and bind to specific receptors on or in target cells

During the reverse anabolic processes

the reduced coenzymes (such as NADH and NADPH) donate their electrons in the biosynthesis of fatty acids, for example. Enzymes called reductases catalyze these reactions

very important facet of signal transduction is that in this cascade mechanism

the signal is also amplified because one molecule of epinephrine activates adenylate cyclase to synthesize many molecules of cAMP, which then activate many protein kinase a enzymes, which then each activate many phosphorylase b kinase enzymes and so on and so on until many glycogen phosphorylase enzymes have been activated within the single cell from relatively fewer epinephrine molecules binding to their receptor.