The Science of Everything Podcast - Episode 75: Cellular Respiration

Episode Date: May 6, 2016

An overview of the processes involved in the generation of energy in cells, including a discussion of the role of ATP in energy storage, glycolysis, the krebs cycle, oxidative phosphorylation, and ATP synthase. Recommended pre-listening is Episode 18: Biochemistry Basics.

You're listening to The Science of Everything podcast, episode 75, Cellular Respiration. I'm your host, James Fodor. So it's been a while since the last show, and I'm sorry about that. I have been busy with other commitments, but I am proud to announce that the show is going to be coming back. Well, it never really went away, but I'm hoping to record more regularly now. I'm not exactly sure how regularly that will be, but significantly more regularly than it has been, at the very least. So today we're going to kick off again with a discussion of cellular respiration, which is something that I've been wanting to do for a while. So in this episode, we're going to talk about the processes involved in the generation of energy in cells, including a discussion of the role of ATP as an energy storage molecule, and then we'll look at glycolysis, the Krebs cycle, oxidative phosphorylation, and ATP synthase.

Recommended pre-listing is episode 18 biochemistry basics. That one is strongly recommended. Some of the other chemistry and biology episodes might also be useful because I'm going to assume a bit of background knowledge, particularly about certain aspects of biochemistry. So that's recommended background knowledge. All that being said, let's jump in and get started. So cellular respiration, what do we mean by that term?

It's important to bear in mind that there are due. different notions of the word respiration. So here we're not talking about the respiratory system, lungs, inspiration of air and breathing, so forth. What we're talking about here is cellular respiration, so that is respiration on a cellular level, being the set of metabolic reactions and processes that take place within an individual cell to convert biochemical energy from nutrients into forms of energy that the cell can use, store and use. So we're looking at a series of chemical reactions here, particularly with the focus on eukaryotic organisms. So those are basically organisms that are not bacteria. Cellular respiration includes a series of chemical reactions which are called catabolic reactions. Now, this means that large molecules are broken up into smaller molecules, releasing energy in the process.

The counterpart of catabolic reactions are anabolic reactions. You might have heard of anabolic steroids before, while the idea there, is that anabolic steroids promote anabolic reactions, that is, the building up of large complex molecules like protein, muscle mass, and things like that. The building up of larger molecules from smaller ones, which takes energy. Anabolic reactions are essential for building, well, any types of proteins or mass structures that make up a body and allow an animal to move and function. But in order for those to occur, obviously energy is required. You need energy to be able to carry out those reactions and that energy comes from ultimately the food that animals consume, but food,

the energy that is derived from food, needs to be converted to a form that the animals can use. This is the purpose of cellular respiration. So I've been talking about converting energy to a form that cells can use. What am I mean by that exactly? So energy exists in the form of chemical bombs. You've talked about that in the past episodes about chemistry and biochemistry and so on. In order to be accessible to the cell, the chemical bonds need to be essentially of a certain type, such that they can be broken and the energy accessed and used to facilitate chemical reactions.

And not just any old chemical bonds that contain energy will do. They have to be of a form that the cell can use. In particular, it's not very useful for the cell to have very large amounts of energy in a single molecule, because often the cell needs relatively smaller amounts of energy in one, go. They don't need huge amounts. So what's useful for a cell is to have energy in relatively small packages, so to speak, which are readily accessible and can be stored and so on. And the molecule that has been adapted by evolution to, or selected by evolution, I suppose, to serve this purpose, is called ATP, which stands for adenosine triphosphate. I would have spoken about this in

previous episodes about biochemistry and so on. So I don't want to get into too much detail about it now. For our purposes, we just need to understand here that it is a molecule that's based on a DNA nucleotide, but it has an extra phosphate group added to it. Phosphate group is basically just a phosphorus atom with four oxygen surrounding it. Adenosine monophosphate, which is sort of the normal form of a nucleotide base in a, you know, that makes up DNA, only has one phosphate group. adenosine diphosphate, ADP, has two phosphate groups, and ATP, adenisine triphosphate, has three of them. Now, the energy that's stored in ATP is stored in between the chemical bond, between the third and second phosphate groups, essentially.

This bond is a weak chemical bond, which is easily broken, thereby releasing energy, allowing the energy to be used for other purposes. This idea that energy is stored in the form of weak bonds might be a little bit counterintuitive, but if you think about it like this, weak bonds are easily broken or more easily broken, which means that then they can be reformed into stronger bonds, which allow electrons to reach lower energy levels. That's what a strong bond is when the electrons are low energy levels. They're stable. They're hard to break because the electrons reach low energy level that it likes to stay at.

So when you break weak bonds, you can make strong bonds, thereby releasing energy. Of course, if an electron is falling from high potential energy to low potential energy, or releasing energy, and that energy can then be used to fuel, perform cellular functions, releasing energy. Again, the idea here is that this weak bond between the second and the third phosphate groups in the adenosine triphosphate molecule is a very convenient storage of energy, and it's also convenient in terms of the amount of energy that it stores, it's a usable amount, but it's not a huge amount. It wouldn't be very useful for a cell to have to release a large amount of energy at once. Think about an explosion. An explosion is a release of a large amount of energy, but that's not very useful for our purposes. We want small amounts of energy so that we can use them to power our computers and televisions and things like that. We don't want huge amounts released all at once. That's not very useful. So it's the same thing for cells. They don't want to have explosive reactions releasing huge amounts of energy all at once. They want to, they want to, have smaller amounts released over a period of time in ways that are useful to the cell.

And adenosine triphosphate is really good for that. So I'm emphasizing this point to explain why it's so important for the cell to form ATP. It needs ATP to carry out almost all of its enzymatic reactions that it uses to build proteins. And if you study all of these different reactions that carry out for all sorts of purposes, ATP crops up all the time. Many, many reactions require it essentially as a co-fact or co-enzyme, which is necessary to provide energy and allow the reaction to occur. Otherwise, these reactions can't happen. The cell will die, and the organism dies.

So ATP is crucial. Cells need that energy storage, and therefore they need to produce a constant supply of ATP. The cell can't store huge amounts of ATP. It can store reasonable amounts of it. A cell can't store enough to last for, you know, months. I forget exactly how long a cell can store ATP stores, but it's not a huge amount of time, because it uses it so rapidly, essentially, because it's used in so many different types of reactions. So, cells need a constant supply of ATP in order to function properly, and ATP's require energy in order to make.

So, where do the ATPs come from, or more to the point, where does the energy required to make ATP come from? Well, the answer is the process of cellular respiration. That's where the energy comes from. And in particular, there are three main parts to this process of cellular respiration that I'm going to discuss. Glysis, the Krebs cycle, which is also called the citric acid cycle, and oxidative phosphorylation. This is a helpful way of breaking up the processes, because each of these steps sort of emerged evolutionarily, separately from the others, and they've sort of been chained up over evolutionary time to form more and more efficient ways of extracting energy, basically. Now, starting with glycolysis, the evolutionarily oldest pathway that we're going to look at here.

Glycolysis does not require oxygen. So it evolved at a time when oxygen wasn't prevalent in the Earth atmosphere and is found in essentially all organisms. The purpose of glycolysis is essentially to begin with glucose and to break it down into two smaller molecules releasing some energy in the process. That's the basic idea. Now, before I get carried away in talking about that, I should start by talking about glucose. Glucose is a six-carbon sugar molecule, a simple sugar. Go back to the biochemistry basic episodes, if you're not sure what a sugar is. It forms the basis of, well, cellular metabolism, basically.

Many other types of molecules are converted into glucose or fit into the glucose metabolism pathway at some point, at least, in order to be broken down by cells. So, so sort of, glucose is the primary energy source of cells. They don't use glucose directly to power biochemical reactions. That will be providing too much energy. Again, it would be like that explosion going off. It's too much energy at once. A single glucose molecule contains too much energy to be useful for most biochemical reactions.

What the cell only needs to do is break it down, store that energy into smaller packages, which you can also store more easily and make more ready use of. For all these reasons, glucose itself, although it's the ultimate fuel source, it's not good as an energy storage molecule or as a medium, so that's why the cell needs to convert the glucose into ATP. And that's essentially the, you can think of as the purpose of cellular metabolism. To start with this six carbon molecule glucose, which has a lot of energy packed in there, and to convert it into essentially as much ATP as we can. That's what we're aiming for here. And the process of cellular respiration

is the process of chemical reactions that brings that about. And what we're trying to understand here is how that happens. So, again, to begin, we start with glycolysis, which is breaking the initial stages of breaking down that glucose molecule from the six carbons, which is what you start with, and the single glucose, into two three carbon molecules, which are called pyruvate. So the process of glycolysis takes place in a series of reactions. There are actually ten chemical reactions, each of which has its own substrate, so that's the starting point, basically, beginning with glucose and ending with pyruvate, and also its appropriate enzyme. So each of these reactions is catalyzed by a different enzyme. I'm not going to go through all of the reactions and enzymes. That's not relevant for our purposes here.

It's too much detail. All we need to understand is that there are 10 different reactions, each following the other, in a linear process, each with its own unique enzyme and so forth. And as a result of this process, we begin with glucose and break it apart into two different, so the reaction splits into two, sort of forks at one point, halfway through.

into two different molecules of pyruvate, which has three carbons. So basically we've chopped the glucose in half and rearranged some stuff, is a simple way of thinking about that. In the process, we release some energy. We produced two ATP molecules. Remember that that was our goal to produce some ATP, so we got two of those. We also produced some water and some heat.

There's two other molecules that this process produces, which we need to talk about. These are called NADH. I won't say what that stands for, because it kind of doesn't matter for our purposes, explain, nor will I explain the structure, because again, it doesn't really matter. The point is that this NADH is another type of energy intermediate molecule. It's not as good as ATP in terms of being useful for direct cellular functions. However, it's sort of a stepping stone.

One NADH molecule carries more energy than a single ATP, but less than a single glucose. So if you want to think of it this way, we start with glucose, which has too much energy and it's too big. We need to break it down into ATP. that's where we're going, that's the aim. NADH is sort of like a middle way between those two. It's part of the way down from glucose, but not all the way to ATP, again,

in terms of the energy that it carries. So the process of glycolysis breaks up the glucose, six carbons into two pyruvate of three carbon each, plus we've got two ATP's out of that, and two NADH. I should say the NADH, the H on that stands for hydrogen, because there's a hydrogen that's attached to the,

the NAD structure, which is the form where the proton, that extra hydrogen is cleave. And that's important because this process of going from the NAD plus form to the NADH form with the hydrogen is important for cellular respiration. And I'll explain that a bit more in a moment. So what have we got so far? We've gone through glycolysis. We've chopped the glucosin 2. Now we've got two molecules of pyruvate plus an ADH plus some ATP.

Where to from there? There's a couple of problems with what we've explained so far. So first of all, in order for glycolysis to occur, we need to start with two NAD plus molecules. Remember I mentioned those just before, that that's the form without the proton added to it, without the hydrogen. We need two of those in order to oxidize the glucose. But the cell doesn't have an infinite supply of these. They're used up in the process of their reaction. We need to find a way of replenishing the supply of NAD plus molecules.

otherwise that they'll be used up and glycolysis would stop. So we need to find some way of getting back those NAD plus molecules. And really the only way of doing that is to oxidize the NADH back to NAD plus. So basically remember, glycolysis takes NAD plus and converts it to NADH. It adds the hydrogen on, essentially. Well, I keep calling it a proton or hydrogen immediately. It's a proton with two electrons, so it's neutralizing out that positive charge. But anyway, the point is that we need to find a way of the hydrogen.

undoing what glycolysis has done to that N80 plus molecule. But how can we do that? I mean, glycolysis just reduced that N80 plus molecule, gave it a hydrogen. Remember, reduced means you add protons to it essentially. If you want to oxidize it back, well, I mean, you could try running glycolysis in reverse, but that would defeat the whole purpose. So that's not really going to work. One way we could do this is to simply have pyruvate. Remember, that's the three carbon molecule that we produced, do the oxidation. So here's the idea. We've got pyruvate, this three-carbon molecule. Have it grabbed that proton back off N-A-D-H so that it converts back to N-A-D-plus, then we've got that N-A-D-plus so that we can go back and feed it back into

glycolycolysis and keep going there. In the process, we can convert pyruvate into lactate, or lactic acid. This process called lactic acid fermentation is carried out by bacteria. It's also carried out by humans when we don't have access to enough oxygen. So if you run, carry out any strenuous exercise, you eventually consume oxygen faster than you can replenish it to your cells in your body, which is why you start breathing heavily and your heart beats faster. But once that occurs, your cells, your muscle cells, which need to contract, they need to use energy to do that, don't have enough oxygen to carry out the full process of southerly respiration, so they have to cut it short. Essentially, they have to produce, they have to get back

those NAD plus molecules faster than they can do it by using oxygen. I haven't explained how that's done yet, but we'll get to that. But there's another way of doing it that replies oxygen. But when your cells can't do that, when there's not enough oxygen to do it, they use the lactic acid fermentation approach. They produce lactic acid. And this is what leads to muscle cramps and other things like that when you do a lot of exercise. Essentially, it's this buildup of lactic acid, which is a byproduct, the anaerobic means of converting those NADH molecules back to NAD plus, and so the process of glycols is going to occur. Now, this might seem like a really good solution, right?

We have this pyruvate thing. I mean, it hasn't really done anything useful for us. We need to find a way of oxidizing NADH back to N80+, so why not just use the pyruvate to do that? We get our NAD plus back, so glycolysis can continue. We get rid of pyruvate to lactic acid. The body can clean that up later. What's the problem?

I think this is a perfect solution. Oh, plus we don't need oxygen for it. You know, having half oxygen rounds kind of a pain, right? You have to breathe and all this stuff. So what's the deal here? Why do we even need to go any further? Well, there's one problem with this method, lactic acid fermentation. Or I should say, before I go on, other organisms, instead of converting to lactic acid, they convert the pyruvate to ethanol.

So some yeast is effectively used to produce bread and alcohol and things like that. Lactic acid fermentation is used to produce things like yogurt. So these processes have a lot of applications in food science. That's parenthetically. What's the problem with this process, be it ethanol fermentation, lactic acid fermentation, whatever. The problem is that the glycolysis plus the oxidation of pyruvate only produces two ATP molecules per glucose.

That's only about 5% of glucose's energy potential. You can get way more ATP out of glucose if you sort of finish the job. Essentially what happens is if you use pyruvate to oxidize the NADH back to NAD plus, producing ethanol or lactate, you leave almost all of the energy of the original glucose in the ethanol or the lactate. It's just left there. It's not accessed. The cell doesn't convert it to useful forms of energy. And that's not ideal because the whole point was to extract as much energy from that glucose as we could. But following this process, we only use 5% of it. What about the other 95%?

Well, it's still sitting there in the form of those relatively high energy electrons in the pyruvate or in the ethanol. And you can see this. If you, you know, ethanol is flammable, you can burn it. That tells you that there is a lot of energy still in there that hasn't been accessed yet. So this is the fundamental problem with anaerobic approaches to regenerating the NAD plus and to dealing with pyruvate. Is that, yep, you can do them without needing oxygen, but they just don't produce nearly as much energy. They only convert about 5% of the energy contained in glucose into usable forms in depth. into ATP. And that's just a big waste, essentially. So evolution doesn't like to be, to waste so much energy like that. So it came up with a solution that is able to access a much larger proportion of the energy of the glucose. And this is what we call aerobic respiration,

which requires oxygen. So in this form, which is the form that, you know, humans and many other animals carry out most of the time when, you know, when we have, we have access to enough oxygen. Under aerobic respiration, pyruvate is oxidized to a different molecule, called Acetyl-CoA, that's COA, which stands for something. Again, we don't care what it stands for. Now, remember, pyruvate contains three carbons. Asetal co-only contains two carbons. The other carbon is released as carbon dioxide. This is the first of the carbon dioxide molecules, which is produced in the process of cellular respiration, and that's why we breathe out carbon dioxide. It's produced as a result of these chemical reactions. Okay, so we go from having this three-carbon pyruvate to this

two-carbon acetyl coa. What's the advantage of that? Well, by itself, this further oxidation produces another two molecules of NADH. Remember, this is another high-energy intermediate, which we can use to make into ATP. We'll get to that process in a moment. So now we go from having 2 ATP plus 2NADH in glycolysis to having, well, still 2 ATP, but now for NEDH. So we've doubled our yield of NADH, so that's already a significant improvement. But that's not really the main benefit of this process, because the real point is now we have acetylcoa. We can feed this into a new reaction series or cycle called the Krebs cycle or the citric acid cycle.

And this is really the key to, well, one of the keys, to understanding how the cell extracts that extra 95% of energy from the glucose. So the Krebs cycle, which is evolutionarily more real, recent than glycolysis occurs inside the inner membrane of the mitochondria. So you might remember these as the energy factories of cells. Bacteria don't have mitochondria. In fact, it's thought that mitochondria actually originally were bacteria in and of themselves, as in separate organisms, and they sort of merged with or were consumed by and became part of other organisms to form the earliest eukaryotic organisms in a process called endosymbiosis. But

That's another issue. What's relevant here is simply the fact that glycolysis, remember the first stage in the process of cellular respiration, that occurs in the cytoplasm. That's just basically the solution that forms the inside of the cell. The mitochondria, though, is a particular subcellular organelle. And so it has its own membrane inside of the mitochondria, and it's inside here, in the inner membrane of the mitochondria, that all of the subsequent processes occur. So the oxidative decarboxylation of pyruvate into acetal coa, the Krebs cycle, and then later on when we get to talking about oxidate phospholation. All these are now happening inside the inner membrane of the mitochondria, no longer in the cytoplasm. So we've moved into a sort of special compartment where this stuff happens. So far, remember, we've taken that three carbon molecule, the pyruvate. We've oxidized it further into acetal coa, which has only two carbons now. One of those carbons is shot out as carbon.

dioxide, and we produce a bit more NADH as a result of that, but we've still got these two carbons left, and there's still a bunch of energy left here that we want to extract. So this is what the Krebs cycle or the citric acid cycle is for. How does it work? Well, it's called a cycle, because unlike Lycolysis, which is linear, the citric acid cycle is actually, well, it's cyclic. Basically, it starts with a particular substrate, which essentially binds to acetalkoa, and then in a series of reactions, this substrate is broken down. so that it reforms the original substrate without the acetal coa. So you can basically think of it as a sort of circular manufacturing process.

You start with something, you add the acetalcoa to it, and then you do a bunch of things to it, a bunch of chemical reactions, spit out some carbons and some ATP and these other things. And when you get back, and by the time you're finished, you end up with what you started with, and so you just add another acetal coer, and you go around again. You spit out more carbon dioxide, you spit out more ATP and other stuff. And you get back to what you started,

you add another astile co-and you keep going around. So this is why it's cyclic. There are eight steps in this process, again, each with their own substrates and different enzymes and co-enzymes and so on. I'm not going to go into all the details of that. Just understand that there are multiple steps in this process with different enzymes, all happening inside the inner membrane of the mitochondria now. And in this process of going around the cycle,

the two carbons in the acetal coer molecule are consumed and essentially released as carbon dioxide. So that's the ultimate fate of all of the carbons in the glucose. Remember, we start with six, and all of them are eventually converted into carbon dioxide. Two of them during the process of oxidative decarboxillation of the pyruvate to the astile colour, and four of them around the Krebs cycle. In the process of the Krebs cycle, we also produce, and this is what we're really interested in, six NAD plus molecules, two FAD molecules, and two ATP molecules.

Now, what's FAD, you might be asking? Well, it's essentially like NAD, except different. It's another one of these high-energy intermediates. It's actually between NAD and ATP, so it carries more energy than an ATP, but less than an NAD plus. That's all we need to know about it for our purposes. Here, the structure and so on is not that important. As a result of glycolysis, then oxidative via coboxylation of the pyruvate, then the Krebs cycle, we've completely ripped apart, essentially, our original glucose molecule. All those carbons are gone and hydrogens and so on. It doesn't exist anymore.

They've been converted into carbon dioxide, basically. Where's the energy gone? Because six carbon dioxide molecules contain much less energy than one glucose molecule. Well, the energy has gone into producing in glycolysis 2 ATP plus 2 more ATP in the Krebs cycle, so 4 ATP in total. And, additionally, 2.8. NAD plus in glycolysis plus two NAD plus in the formation of pyruvate to estalcoa plus plus in the CREB cycle, making a total of 10 NAD plus molecules plus the two FAD that I mentioned.

Because what are we going to do with these things? Well, before I explain that, there's one little extra technicality that I haven't mentioned. Actually, there are quite a few technicalities that I'm glossing over here, and a few things I'm simplifying or... but these processes are very complicated. So I am simplifying, and there are a few details you might read that are slightly different, but this is an introductory episode, so I don't think that's too problematic. I'm trying to get you to understand the main ideas, not get bogged down into all these little details. But there is a detail that is important, I think,

because I said that glycolysis, remember the first 10-stage linear process, breaking down the 6-carbon glucose into the 3-carbon, 2 lots of 3-carbon pyruvate, that produced 2-N-A-D-plus molecules. But I said that this glycolysis occurs in the cytosol. But then the Krebs cycle and all the other things occur inside the mitochondria and inner membrane. We get eight additional NAD plus molecules there. How do the NAD plus molecules produced in glycolysis get into the membrane? You might be wondering.

Well, if you hadn't wondered this, you should have because that's actually important. These NAD plus molecules need to be inside the mitochondria in a membrane in order for them to take, in order for them to be utilized, the energy extracted to the final form and converted to ATP. In fact, what happens here, as I understand it, there are actually a number of different ways this can occur, but one of the mechanisms is essentially that the NAD plus, excuse me, the NADH molecules,

that's the energetic form, the NAD plus is the starting form, the oxidized form, which is then reduced, gains energy essentially, to form NADH. Anyway, the NADH doesn't actually move into the inner membrane. It actually stays in the cytoplasm. What it does is it passes its energy on by converting an FAD into an FADH2 molecule. So remember these FAD molecules I mentioned before, that two of these are produced in the Krebs cycle. Well, it turns out we actually get two more, and these come from the NADH molecules that are produced from glycolysis.

FAD molecules do have, or FADH2, do have somewhat less energy than the NADH, but they still have more than ATP. So that's one way that you can get these NADH molecules containing this energy that is outside the mitochondria inner membrane and get them where we need inside the inner membrane. Essentially you convert them by a membrane bound protein or enzyme. You convert the NADH into an FADH 2, which is again this other intermediate. So with that little technicality, what we've got so far are 8 NADH molecules waiting for their energy to be extracted and converted to ATP, which is what we really want, plus 4 FADH 2 molecules, which also are waiting for their energy to be extracted. So essentially 12 high energy intermediates. We've got 8 NADH 4FADH 2.

Alright, so how do we extract this energy? The answer essentially is oxidative phosphorylation. This is the last of the three main stages or phases in cellular respiration that I mentioned. So oxidative phosphorylation goes after the Krebs cycle, and it's essentially the process of extracting the energy from these F-A-D-H-2 molecules and N-A-D-H molecules, and converting it into a form that's used to make ATP. Now, this is actually, well, it's probably the most elegant and fascinating of the stages in cellular respiration, because unlike in the previous reactions, where essentially ATP or N-A-D-H-H, or whatever is made directly as a result of the enzymatic reaction. In the case of oxidative phosphorylation, that doesn't occur. FADH and NADH don't directly make any ATP. What happens is that they, the NADH and the FADH2, are both oxidized back into their low-energy versions, the FAD and the NAD plus.

So they lose those hydrogens, essentially. and in doing so, they lose energy, and that energy goes into producing a proton gradient over the membrane. Remember, this is all happening inside the inner membrane of the mitochondria? So there's an outside to that membrane, which is on the other side of the membrane. Oxidated phosphorylation essentially is the process of pumping protons from the inside of the mitochondrian inner membrane to the outside of that membrane. Why would you want to do such a strange thing? I thought we were trying to make a

ATP, right? Well, the idea here is that if you can create a proton gradient, you can use this gradient as a source of energy, essentially. A proton gradient is a source of energy in two ways. First of all, it's a concentration gradient. From chemistry that we've talked about before, we know that if you have any chemical species in higher concentration on one side of a membrane over another, a semi-permeable membrane, then through osmosis, we'll have those molecules or particles or whatever they are, move from the area of high concentration to low concentration, and in that directional motion we can extract energy. So that's a source of energy, essentially. So basically protons flowing across the membrane. They'll flow from high concentration to low

concentration. That will be their tendency anyway, if they can, if we allow them to flow across. The other way that a proton gradient is a source of energy is because protons obviously are electrically charged. So if we have a bunch of them on one side of the membrane and no offsetting electrons there, then we get a charge differential across the membrane. And again, if you have a region of positive charge, then those will tend to repulse and move away to regions of negative charge to neutralize. So there's two reasons why a proton gradients useful, essentially, because of the chemical concentration and also the chemical charge different, sorry, the electric charge differential. Both reasons mean that this region of high concentration of protons is essentially energy rich. We can use it to produce energy. So that's why we might want to produce this proton gradient. But how does it happen? I mean, how do we go from FADH and NADH molecules to a proton gradient? Well, essentially how it works is that there is a series of four complexes, a protein complexes that span the membrane between the inside and the outside of the mitochondria in a region.

These proteins essentially stick out either side of the membrane. And on the inside of the mitochondrial membrane, the NADH and FADH2 molecules interact with the proteins, essentially giving up protons and high-energy electrons to the proteins that are in the membrane, to these complexes, and then the complexes pass the electrons along in the process using the energy from the electrons to pump protons across the membrane. So these complexes which are in the membrane, they're studded in the membrane, they're numbered one through four in accordance with the order that the electrons move through them. These complexes form what we call an electron transport chain, because literally it's like a chain. One complex hands the electrons to the next one, which hands this to the next one, but hands us to the next one. That's helpful because the electrons start off in a high energy state, and as they move across the chain, they gradually lose energy. And this comes back to the thing we mentioned before. The whole point is it has to happen gradually.

We don't want the electrons to suddenly lose all their energy. It falls straight from the top floor of the building down to the bottom because that will not allow us to store that energy in a useful form. We need it to release it slowly so that we can extract it and then store it in the ATP molecules. So that's why we have this chain that allows us to gradually extract that energy from these high-energy electrons, use it via essentially conformational changes of the proteins, to pump protons from the inside to the outside of the membrane, producing a proton gradient

over the other side of the membrane. In the process, of course, the NADH and the FADH2 molecules are oxidized back into their low-energy forms, essentially, the NAD plus and F-A-D. So they're ready to then go back and be involved in the reactions again. Remember that we needed those forms to start to get the process going in the first place. So we need to regenerate those low energy forms. So we've got the protons off those, that's good, and we've extracted the high energy electrons that come with the protons, and then we've extracted the energy from those high energy electrons and use that to pump protons over, and now the energy is sitting in the form of a proton gradient across this mitochondrial membrane. There's a couple of issues here.

First, we need to figure out a way of converting this proton gradient energy into ATP, which is ultimately, again, what I'm interested in. The second issue is, is we need somewhere to put all these electrons. They're not high-energy electrons anymore. They're kind of lower-energy electrons because we've extracted all their energy through this electron transport chain. So we can't give them back to a molecule like N-A-D

plus or F-A-D, for example, because essentially they would need to be much higher energy to be accepted by these molecules. So we need a molecule which has a really great affinity for electrons. That is, it will accept even very low-energy electrons. A different way of thinking about this. is that you can think of different atoms or different molecules as being

different depressions in the ground, the depressions, the sort of depth of the depression representing how much of a pull for electrons it has. The deeper the depression, the greater the pull for electrons it has, the high electronegativity. You can think about it, although we're talking about a molecule here, not an individual atoms, so it's a bit of an analogy. But that's the essential idea. Some molecules slash atoms have much greater pulling power for electrons than others. In order to accept a very low-energy electron, we're going to need one that has a very great pulling power,

because the others won't, essentially, won't have the pull needed to attract and hold that electron. What is a atom-slash-molecule that has very high electronegativity? Well, oxygen, actually, if you look at your peer at a table, I'll go back to one of the chemistry episodes where we talked about this. Oxygen is, I think, the second most electronegative element, and molecular oxygen, two oxygen atoms bound to each other, has a very great affinity for electrons. So, turns out this is the perfect substrate,

to act as the final electron acceptor. So it accepts these now low-energy electrons, grabs them and holds onto them, so they fall down into essentially a very low-energy state now. They're not going to come away from those oxygens, or at least you're going to have to add a lot of energy to pull them out of that again, so it's quite difficult. This is what makes oxygen such a great terminal electron acceptor. But they don't stay in the form of oxygen. They react with grabbing some extra electrons, so they need to balance out that charge now, so they react with some protons to form water. Of course, if you take oxygen and you add hydrogen to it, or H you get in the right ratio, you get water, H2O. So that's the fundamental idea here. We take oxygen, use it to soak up

these extra low-energy electrons that we need to put somewhere. Obviously, they can't build up, otherwise there'll be a negative charge, and that's going to cause problems if we allow that to build up. So we soak up those electrons with oxygen, balance out the charge by adding some protons, and if you add those in the right ratio, hey, you get water. That's why we produce water as part of this process. So we've solved that problem, we've soaked up those electrons. One problem remains, how do we extract the energy from this proton gradient and turn it is ATP? Well, that's where the last member of the oxidative phosphorylation electron transport chain,

although sometimes it's considered separate to the chain, but whatever. For our purposes, we can think of it as the fifth member of this chain. That's where it comes into the picture. called ATP synthase. So as the name indicates, it actually makes ATP. It takes as substrates adenosine diphosphate, so that's with two phosphate groups bound to it, and also inorganic phosphate, which is just the one phosphate group, and also energy. And then it binds them together, so you produce adenosine triphosphate, so three phosphate groups. With, again, the bond between the second and third phosphate groups being relatively weak, which means it's a high-energy bond,

There's energy to extract there, which then can be used for processes of metabolic reactions elsewhere in the cell. That's the whole point of our exercise is to produce these ATP molecules. And the proton gradient is used by ATP synthase to do exactly that. But how does it work? Well, I'm not going to talk into about the exact details of how ATP synthase works. In fact, it's only quite recently been discovered, and I think we're still learning the exact ins and outs. but it's quite an intricate mechanism. Essentially, it has a rotor.

The whole molecule, well, part of the molecule, some of the subdomates, rotate around essentially. The basic idea of what happens is that, again, ATP synthase extends across the membrane over the mitochondrial organelle membrane, and the protons move through a channel inside the center of the protein, of the protein complex, the ATP synthase complex.

But they don't just move straight through, that wouldn't be very helpful. As they move, they interact with the protein in such a way that it changes its confirmation. And essentially, there's this sort of little mechanism in the middle that rotates around. There are different positions that it can be in. You can think of it as if you're looking at it, if you were looking at ATP synthase, so radially, that is top-down, viewed so that you were looking at it down through the membrane. it's pointing up to me, the channel that the protons travel through is facing me so I can see, I mean, if you can imagine, I could see the other side of the membrane, and then the proton

coming through and out to me. If you look at it from that perspective, it actually looks a bit like a clock face, vaguely, so that there's this sort of central part of the protein complex that rotates around as protons move through the central channel. The purpose of this rotation is that it in turn affects the confirmation of other subdomains of the protein complex, which change their shape, change their confirmation. As they do so, they move through a series of stages where one confirmation allows adenosine diphosphate and inorganic phosphate to bind on to the enzyme, close to each other, and then another confirmation essentially squishes them together.

It catalyzes the reaction so that they go from being separate molecules to being joined together. that catalyzes the formation of a bond. And then in another confirmation, that a proton will open up and the ATP will be allowed to leave. It'll diffuse away. That's a very sort of rough explanation. But the basic idea is that it's this little factory mechanism as it rotates around.

It brings in the ADDP and phosphate. It changes confirmation, smushes them together, and then changes confirmation again, and it allows them to diffuse away. And there are actually three different sites around the edges of the ATP synthase molecule where this is happening at the same time. at different phases. Each is in a different stage, but it's all happening at the same time.

The key point to understand, though, is that through very clever, conformational changes and very intricate, evolutionarily derived internal mechanisms of the ATP Synthes Complex, it is able to use that motion of the protons moving through its central channel to catalyze the formation of bonds between the ADP and the inorganic phosphate to form ATP. And, of course, in the process, we have the proton coming across the membrane, and so equalizing the charge. Obviously, if we had indefinite buildup of proton gradient over the other side of the membrane, and that would also be bad, that would not be sustainable in the long term. What happens is we only have a fairly small build-up, it's gradually being, well, there's a dynamic equilibrium because we're pumping protons over there through the oxidative phosphorylation,

but then we're bringing them back down through ATP synthase, and so the net charge on the other side is kept at a sustainable level. So that essentially rounds out the process. each NADH molecule that we oxidize back to NAD plus is able to pump 10 protons across the membrane. Each FADH2 molecule that we oxidize is able to only pump six. So remember I said that FADH2 has less energy than the NADH. That's shown by the fact that NADH pumps 10 protons per one of those molecules, whereas FADH only six. and it takes about four protons to produce,

across, coming down through the ATP synthase, to produce one molecule of ATP, which means that given NADH is able to produce, is able to pump 10 protons, and then you need four, for one ATP, it's about 2.5 ATP are produced for every NADH molecule, and about 1.5 for every FADH2 molecule. You'll notice these are not stochymetric ratios, that is, they're not whole numbers like we have for most chemical reactions,

that's because this process is not a sort of standard chemical reaction. It's a chemio-osmosis, the process is called, where we have this gradient, proton gradient, traveling across the membrane. It was not what researchers were expecting when they were originally studying this. It makes it hard to give exact numbers. You'll see slightly different numbers for the yield, the ATP yield, it's called, of this process as a result, because it does depend on certain assumptions you make

and the measurements vary a bit. That detail need and concern is too much here. So let's zoom out again and review and summarize the whole process and also count up and sort of tally where we're

getting the energy from and look at our final yield of ATP. So remember, we started with a single molecule of glucose which contains six carbon atoms. The carbon hydrogen bonds is fundamentally where the energy is coming from. Those carbon hydrogen

bonds are relatively high energy bonds, that means that the electrons are in a relatively high energy state. In order to extract that energy, we have to move those electrons to a lower energy state. And ultimately, that's going to be in the form of carbon oxygen bonds in the carbon dioxide and even more importantly, oxygen-hydrogen bonds in the form of water. So that's the overall process here, will be to convert glucose plus oxygen into water plus carbon dioxide. And that's the overall chemical reaction for respiration, which you may have seen before. Essentially, that's what we're doing.

We're taking those high-energy electrons from carbon-hydrogen bonds and converting them into low-energy electrons in carbon-oxygen and oxygen-hydrogen bonds. The purpose of that, of course, if we can convert high-energy electrons into low-energy electrons, we can extract the energy and use it to convert ADP into ATP, which is, of course, the energy currency of cells, as I've talked about a lot today. So that's our goal. How do we do it? Well, the first step is to take the six-carbon glucose molecule, break it into two through glycolysis, this linear process of 10 reactions. In the process there, we break up the glucose into two pyruvate molecules, and we extract two molecules of ATP, plus two molecules of NADH. Glycolycolysis occurs in the cytoplasm, so we need to get these NADH molecules into the mitochondria in the membrane for the subsequent processes. Essentially the way we do that is a coupled reaction using membrane-bound enzymes,

which essentially take the energy from our NADH molecules and convert it to, and pass it on to FADH molecule, FADH2 molecules which are inside the membrane. We've now got, practically speaking, two ATP molecules and two FADH2 molecules inside the inner membrane where we need them. But we've also got a build-up of NADH, which is a problem because we need NAD plus in order for glycolycolysis to occur. As I've been saying, the glycolysis and many of the other reactions that we discuss in the process of cellular respiration are oxidation reactions. That is, the substrates lose electrons. Remember, the whole point is we're trying to pull out these high-energy electrons and extract their energy.

So, of course, the substrates are going to have to lose electrons in order that to happen. That means we need something to grab the electrons, so we need something that is, if whenever something, something is oxidized, something else has to be reduced. So what is reduced are these high-energy intermediates, or low-energy intermediates then become high-energy intermediates. In particular, the NAD plus. That is reduced to form NADH. Remember reduction is when you gain electrons,

or also when you add hydrogens, which come along with electrons, that's sort of the biochemistry way of thinking about reduction. NAD plus to NADH is a reduction reaction. Now, we need to undo that. We need to oxidize NADH because we need to have a store a pool that's replenished of these NAD plus molecules. Otherwise, glycolysis can't occur. So how do we do it?

How do we replenish the pool? How do we re-oxidize the NAD plus molecules? Well, the one route is to simply use pyruvate to do that, to convert it to lactic acid or possibly ethanol. That replenishes the NAD plus molecules, and it doesn't require any oxygen. downside, we can't, it locks up all of the rest of the energy that is still contained in the lactate or in the ethanol and doesn't allow the cell to access it. So we only get that yield of two ATP molecules, 5% of the total energy potential, not very useful. So we only do that, we being, you know, you carry out when we can't get access to oxygen. And even that it's not sustainable, it just can carry you on in the intermediate time until you can get oxygen back again.

No. What we need is a further process in order to extract the rest of that energy. And that occurs in the next two parts of cellar respiration, the Krebs cycle and oxidative phosphorylation, both of which take place, as I mentioned, inside the inner membrane of the mitochondria. So in the Krebs cycle, this is a cyclic process which involves eight reactions. Before we get to the Krebs cycle, we have to grab our pyruvate, whack off a carbon, which is then converted into carbon dioxide. We've now gone from three carbon pyruvate to only two carbon molecule, which is called acetalcoa.

That acetalcoa is fed into the Krebs cycle, in the process of which we go sort of round that cycle, wind the crank around if you like. We completely break up the acetalcoa, spit out two more carbon dioxide molecules, so now we've completely broken up and destroyed essentially the original glucose molecule. All of the carbon is gone. But importantly, we extract the energy,

We extract those high energy electrons by oxidizing 6 NAD plus molecules and 2 FAD molecules plus producing another 2 ATP molecules So we've now got a total of 4 ATP molecules and we've totally broken up our initial product the glucose But we have that problem of what do we do with these 8 NAD H high energy intermediates and plus 4 FAD H2 Slightly less high energy intermediates we need to convert them into a ATP. They can't stay as they are because that's not useful for the cell. This is where oxidative phosphorylation comes into the rescue. Oxidative phosphorylation is the process by

which these high-energy intermediates pass along their high-energy electrons to an electron transport chain of proteins which are embedded in the mitochondria in a membrane. They pass along the high-energy electrons which gradually move across the chain, moving from one protein complex to another. As they do so, they gradually move from a state of being high-energy to low-energy, with the energy used to pump protons across from the inner to the outer regions of the membrane, producing a proton gradient. Each NADH molecule is able to pump 10 protons across the membrane in this manner. Each FADH2 molecule able to pump six protons in this manner.

So this is how we use up all those high-energy intermediates. We oxidize them back to their original forms, which, again, we need to keep the cycle going. That solves that problem. But now we have a proton gradient and we need to find a way of converting the proton energy, chemical and electric potential energy, into ATP, which is what we want. That's where the last enzyme complex comes in, ATP synthase. ATP synthase allows the protons to travel back down along the membrane through an interior channel, and in doing so extracts the energy that's released by this falling proton, essentially falling down a chemical gradient, an electric gradient, uses it to synthesize. synthesize the formation of ATP from ADP and the inorganic phosphate. It takes four protons to produce one ATP molecule, so that means each of the NADHs produces about 2.5 ATPs,

and each FADH2 produces about 1.5 ATPs. So if you do up the tally with the 4 ATP that we originally produced, plus 2.5 times the 8 NADH, plus 1.5 times the 8 NADH, plus 1.5 times the 3.5 times the the 4 FADH 2, converting the high-energy intermediates through the oxidative phosphorylation process, we come to a total of about 30 ATP. As I said, you might find different numbers in different places because there's slightly different ways to do the accounting, and it depends on various factors, like the process used to transfer NADH inside the mitochondrial membrane, but those details won't concern us here. The basic point is that instead of only

producing 2 ATP, as occurred, in the case where we... just used pyruvate to re-oxidise NADH back to N80+, locking up the 95% of the energy in an inaccessible form in the lactic acid or ethanol. Now, by using the process of the Krebs cycle, oxidative phosphorylation, ADB synthase, we are able to extract the full amount of the energy, obviously not 100%, literally, because, you know, second law of thermodynamics, you can't have a fully efficient heat agent, but, you know, a much, much larger proportion of the total energy available in glucose is used to produce. a grand total of about 30 ATP molecules, which are then used to provide fuel for the cell. So that brings us to a conclusion. That's the core concepts that I wanted to get out about

the cellular respiration process. Hopefully it was somewhat clear. Again, diagrams are really helpful for this. I'll post them up on the Facebook. If you enjoyed the podcast and would like to send me some feedback, you can contact me at FOD12.g.com. That's FOD. s12 at gmail.com. Thanks for listening and I'll talk to you next time.