#29 Biochemistry Citric Acid Cycle Ii Lecture for Kevin Ahern's Bb 451/551

Kevin Ahern: This is really bad. I was not fishing for that. The second is some of you have noted that there are some discrepancies between what I talk about with E1, E2, and E3, and what the book says. The book is actually more precise than I am, and I'm trying to simplify this for you. E1, E2, and E3. So don't sweat the discrepancies between the two. I'm really not that interested in you just dissecting what's E1, what's E2, what's E3. It's the big picture about what's happening in this. You should know the process but remember that these are happening between subunits and so as things are happening between subunits, it's really hard to say this is happening only here. So don't sweat the discrepancies that are between what I said and what the book says. That said, let's go back into the citric acid cycle. And we'll actually finish it up. We're in very good shape for stuff. Alright, and we shall move forward. So last time when I talked, I showed you this reaction here, which was the alpha ketoglutarate reaction at the very end. And I said that this is a second oxidative decarboxylation. I pointed out that it was catalyzed by the enzyme alpha ketoglutarate dehydrogenase and that that enzyme had a structure and a mechanism that was essentially the same as the mechanism that was used by the pyruvate dehydrogenase complex. The same coenzymes. The same basic mechanism. One slight difference in mechanism is that the bacteria and yeast don't subvert this one. So they can subvert the other one, go through decarboxylation but not oxidation. That's what gave acid aldehyde which gave ethanol. They go all the way through this reaction. So they don't cut it short, alright. Other than that, mechanistically, this reaction is essentially the same as the pyruvate dehydrogenase complex. You notice that by this point we've gotten to an intermediate that has four carbons. In addition, we've created an intermediate that has a high energy bond. That sulfur bond between the carbonyl group right there and the CoA is a high energy bond, and we'll see that that bond will come into play in the next reaction. So this high energy bond is very, very useful for the cell, and what it's done is the cell has used the energy of oxidation to help drive the formation of this high energy bond. Well, the next reaction is a reaction that seems on the surface to be catalyzed by an enzyme that is misnamed. So the reaction, let's look at it first. The reaction is shown right here, Succinyl CoA, plus inorganic phosphate, and GDP gives us succinate, succinyl CoA, and GTP. Now I think last time I actually described this guy as an activated intermediate. From the definition I've given you, technically it's not, but we'll treat it as if it is. An activated intermediate has a high energy bond and uses the energy of that bond to donate a part of itself to something else. Well, there's no part of itself here that's being donated over here to make something else. But, this high energy is used to drive the formation of GTP. So we'll treat it as if it's an activated intermediate even though, technically, it's not. Notice we're making GTP instead of ATP. Some organisms make ATP, not GTP. We're not gonna sort through that. We'll treat it as if it's just GTP that's being produced. GTP, you should recall from last term, has the same energy level as ATP does. Alright, exact same energy level. So we made a high energy triphosphate just as if we were making ATP. Now what was confusing about this is the name of the enzyme. The name of the enzyme is succinyl CoA synthetase. What does the name of that enzyme tell you it's doing? It's making succinyl CoA. What's wrong with this process? Well, this is breaking down succinyl CoA, right? Why do we give this enzyme this idiot name? Student: It's succinyl CoA synthetase? Kevin Ahern: Yes. Kevin Ahern: Succinyl CoA synthetase. Why did we give this enzyme this idiot name? Student: Is it reversible? Kevin Ahern: Of course. It's a reversible reaction, and, when people first started studying the enzyme, they were studying the other reaction going the other way. The name stuck. And so we call it succinyl CoA synthetase. But remember this is a reversible reaction. It's essentially any enzyme in a reaction is reversible. This is reversible as well. So, that seems sort of a backwards name, but that's what it does. Succinyl CoA synthetase. Now, we've got a four carbon compound. From this point forward, all that we're gonna do is rearrange that four carbon compound back into oxaloacetate so we get back to our starting point. And to get back to oxaloacetate, we've got a couple of oxidations that we've gotta go through. Alright, now the first of these oxidations... [clearing throat] ...excuse me, is interesting and a little unusual actually. It's this guy right here. Succinate going to fumarate. For some reason, your book groups all three of these on the same slide. Succinate going to fumarate is an oxidation. It's an oxidation like you haven't seen before. It's an oxidation that is simply removing hydrogens and their electrons. We've talked about dehydrogenases, but they weren't technically physically pulling off only hydrogens. That's what's happening here. This pulling off only hydrogens takes a different electron carrier. There are energy reasons for that. I'm not gonna go into why they are there. But FAD is the acceptor of the electrons in this reaction. The enzyme that catalyzes this reaction is known as succinate dehydrogenase. Succinate dehydrogenase. Well, what we're doing in taking away two protons and two electrons is we are making a double bond intermediate. That double bond intermediate is known as fumarate, as you can see here. You'll see fumarate has a trans-bond. And I point these things out to you because this reaction that you see right here, and this reaction that you see right here, and this reaction that you see right here, that is all three of these reactions occur in fatty acid oxidation. All three of them. We will talk separately about fatty acid oxidation, and when we do, I'll remind you of these reactions. But fatty acid oxidation involves dehydrogenation, taking away the hydrogens. It involves adding water. It involves oxidizing the hydroxyl group. So if you learn this set of reactions, you learn the reactions of fatty acid oxidation at the same time. Because they occur in exactly the same sequence, very, very similar mechanisms, and the same electron carriers. So the first enzyme here in this one is succinate dehydrogenase. This enzyme we'll actually see later when we talk about electron transport, because this enzyme actually plays a role in electron transport. This enzyme is the only enzyme of the citric acid cycle that's found in the membrane of the mitochondria. Succinate dehydrogenase is found in the membrane of the mitochondria. It's the only one found in the membrane. All the others are dissolved inside. We're getting close to our final goal. We've got fumarate. We add water across that double bond. We get an OH one, and we get the H on the other side. The enzyme that catalyzes this reaction has a very complicated name. It's known as fumarase. F-u-m-a-r-a-s-e. Malate goes to oxaloacetate in what we will call here the last step of fatty acid synthesis, although we remember it's a cyclic process. What we've done is we've gotten back to our starting point. Now this reaction in going from malate to oxaloacetate is an unusual one. Well, what's unusual about it? Well, it starts with a hydroxyl. It ends with an alpha keto group that you see here. That's not unusual. What's really unusual about this reaction is that it's an oxidation that is not, underline not, very energetically favorable. It's an enzymatic, I'm sorry, it's an oxidation that is not very energetically favorable. All the other reactions I've talked to you are reasonably favorable. A little bit negative. A little bit positive. But basically, they're reasonably favorable. This guy has a fairly positive Delta G0' value. Now when we talked in glycolysis last term about the aldolase reaction, you may recall that the aldolase reaction had a high positive Delta G0'. How did the cell get around that aldolase reaction? Pushing and pulling. It started pulling away product, and it started pushing by making more substrate. Right? Pushing and pulling was the key to how glycolysis in the cell gets around that large Delta G0' value. In this case, we see pulling. And how do we see pulling? Well, pulling you remember, involves the loss of product. And if you recall that very first reaction I gave you, the citrate synthase reaction I said was energetically very favorable. That means that the product, oxaloacetate, is quickly taken away by the citrate synthase reaction. So this reaction actually gets pulled by the citrate synthase reaction. As a consequence of this, the citric acid cycle is energetically very favorable. It goes around quite readily. We start with a four carbon compound. We add two carbons. We lose two carbons. When we come back, we've still got a four carbon compound. Now just anecdotally I will tell you that the two carbons that we add are not lost in that first rounding cycle. It's not until the second round before they start being lost. There are people who get really upset tracking those carbons and so forth. I think it's anecdotal for our purposes, okay, but I'll just mention that to you because some books will make a big point of that when you look in the books. Kevin Ahern: Yes? Student: Did you mention what enzyme catalyzes that? Professor: I'm sorry. I didn't mention that. Thank you for reminding me. The enzyme that catalyzes this is malate dehydrogenase. Professor: Malate dehydrogenase. This is malate, and malate dehydrogenase catalyzes this reaction. Okay, so that's the citric acid cycle. Now we're gonna talk about the citric acid cycle in a lot of ways. The number one thing I want you to be thinking about with the citric acid cycle, I'm gonna come back to this over, and over, and over, is the fact that it's an oxidative pathway. It's one of the cell's most important and most used oxidative pathways. We think back to glycolysis, we only had one oxidation reaction in ten. Here's the citric acid cycle, and we've got eight reactions, four of them are oxidative. Four oxidation reactions here in the citric acid cycle. Because we have four oxidation reactions, we produce four reduced electron carriers. Three of those are NADH, and one of those is FADH2. Now this is not just something that's parenthetic that I'm mentioning about the oxidation. We're gonna see that the oxidation is a very, very big determinant on whether or not the cells run the citric acid cycle. I'm gonna show you that in just a second, but I want to plant that idea in your head. The oxidation that's done in the citric acid cycle requires what? It requires electron carriers. Why did cells in glycolysis go through fermentation? What were they trying to make? Final exam question. They were trying to make *** because they had to keep that one oxidation reaction going in glycolysis, and *** is needed for oxidation. In the citric acid cycle, cells need ***; cells need FAD. And now here's the catch, they don't have fermentation as an option to keep it going. Fermentation is going out in the cytoplasm. It's not occurring in the mitochondrion. So one of the things that's really going to determine whether or not the citric acid cycle occurs is whether or not reduced electron carriers are available. Is *** available? Is FAD available? It turns out that cells cannot move electron carriers across the mitochondrial membrane. You can't say, "We're gonna make a bunch of *** "out here in the cytoplasm. "Oh, let's move it into the mitochondrion." We can't directly do that. There are ways of getting electrons in or out, but they're not simple. Okay, alright. So that's a very, very big thing that we need to understand, alright? And that actually leads me to my next topic, which is... Where'd I put it? No, here, right here. Okay, alright. I'll come back to the other topic there in a second but I want to talk about this because I'm on the topic at the moment, alright? When we think about the citric acid cycle, alright, you think, "Oh boy, here's another cycle "we've got to learn the regulation of." Glycogen metabolism was terrible. Glycolysis, there were three enzymes, okay. And now we've got the citric acid cycle, and we've got to say, "Oh my God, I've got to memorize this. "I've got to memorize this. "I've got to memorize this." I'm gonna keep it really simple because it turns out that most of these allosteric effectors play very, very little role in controlling their enzymes. Very little role. The most important regulation of the citric acid cycle is just what I told you, the availability of *** and FAD. We can look at the reactions of the citric acid cycle and almost completely decide that the whole cycle is going by the availability of those two. In general, when we have plenty of ***, okay, and FAD, the cycle is gonna go. Are there complicating factors? Yeah, a little bit. What's one of them? The only one we're gonna worry about is ATP. And if you look, in every case, what does ATP do? It turns off the enzyme. That makes sense. High energy. Do you want to be running the citric acid cycle? Of course not. For the same reason as I've talked about before, that you don't want to be lighting a fire in your fireplace when it's summer. You got plenty of energy. You don't want to be burning more. You want to stop that process. In general, when cells have plenty of ATP, they also have plenty of NADH. They also have plenty of NADH. So these two usually go hand in hand. When cells have low energy, they have very little ATP, and they have plenty of ***. Now they want the thing to go. So this is very simple. I don't even care if you know this enzyme is regulated or this enzyme is regulated. I think you should know this guy up here is regulated. This is pyruvate dehydrogenase. But all I care about down here that you know is that a lack of *** is gonna stop any of the oxidation reactions. That includes this one, and that includes this one. Anything that has a need for an electron carrier, *** or FAD, that reaction is gonna be stopped if it's not available. Period. Now as I said, I'm gonna come back to that. Because why is that important? The reason that's important is if we understand our bodies and how our bodies are burning energy, we start to understand when the citric acid cycle is running. Now I'll come back and talk to you about this later, but since I'm a little ahead, I'm gonna give you just a sort of a brief preview of that. When cells have plenty of oxygen, we talked last term, what is available? ***, cells need oxygen to make ***. It is when they ran out of oxygen, that's when they started to ferment, and that is when *** concentrations went down. Everybody remember that? Alright, plenty of oxygen. Plenty of ***. Are there limits to that? Yeah, there are. We're not always having a situation where we've got plenty of *** because what's *** converted into? NADH, right? And NADH, we will see, is used to drive electron transport. And electron transport is used to drive the synthesis of ATP. What if we have a lot of ATP? Does this thing start backing up? How would that happen? Let's imagine we're all sitting here drinking beer, eating pizza, and pointing at Kevin Ahern up there giving a lecture. Kevin Ahern is burning a little bit of energy up here in front of you walking back and forth and back and forth. You guys wonder why I pace like this. It's actually to burn off energy. But you're sitting here. You're not burning up much. What's happening to your ATP levels? Are they high, or are they low? Kevin Ahern: Energy levels are high, right? When your ATP levels are high, okay, it turns out that oxidative phosphorylation stops. Oxidative phosphorylation is what produces ATP. When oxidative phosphorylation, and by the way, you don't need to write any of this down. I'm gonna talk about all this later when we talk about oxidative phosphorylation. I just want you thinking about it. Oxidative phosphorylation is stopping. It turns out oxidative phosphorylation and electron transport, one depends on the other. If I stop oxidative phosphorylation, I stop electron transport. And electron transport is necessary for me to make *** from NADH. Too much ATP. That means I'm gonna have a lot of NADH. If I have a lot of NADH, what's gonna happen to the citric acid cycle? Student: It will stop. Kevin Ahern: It's gonna stop. What's gonna to happen to my burning up of things? It's gonna stop or slow down a heck of a lot, right? If it slows down a heck of a lot, what's going to happen with all that food that I'm eating and that beer that I'm drinking? It's not gonna get burned off, right? And what am I gonna start making? Fat. Now we'll see this more at the molecular level when I start talking about oxidative phosphorylation and electron transport, but you can start to see the glimmer about how these pathways, being interconnected like they are and dependent upon each other as they are, are completely controlled by very simple things. Very, very simple things allow us to control those. Questions or comments before I move on? I can tell you to start writing again. So we'll get to that right now. I'll remind you the relationship to glycolysis. The main thing glycolysis was doing was it was giving us a three carbon intermediate, pyruvate. That pyruvate was getting converted into acetyl CoA. And that acetyl CoA was being used in the citric acid cycle to make GTP, NADH, and FADH2. From the cycle, you saw that one acetyl CoA gave three NADHs, it gave one FADH2, and it gave one GTP. How many acetyl CoAs do we get per glucose? We get two, right? Two three carbon intermediates give two two carbon intermediates. So if we started with glucose and we looked at what it produced in the citric acid cycle, we could say that the carbons from glucose are giving us six NADHs, two FADH2s, and two GTPs. Now we'll talk about this more when we talk about the oxidative phosphorylation again later, but I'll just tell you now if we convert those NADHs and FADH2s into ATP, and we convert the GTP into ATP, and we add all these guys up, if we start with one glucose molecule and we count all those ATPs that came during glycolysis, we start with one glucose molecule, we can produce a total of 38 ATPs by using glycolysis and oxidative phosphorylation. So even though glycolysis itself doesn't give us much upfront, it's getting us two NADHs, it's getting us two pyruvates, and it's getting us two ATPs. All of those are important energy considerations. Most importantly, for our purposes here, glycolysis is giving us a three carbon intermediate that feeds into the citric acid cycle. Now I need to introduce another term to you. And I don't do this to complicate the picture. I'm trying to keep the picture simple here, but let me introduce this term because it is important to understand this bigger picture of the citric acid cycle. The citric acid cycle does not exist in our cells solely to make energy. Energy is a very important consideration, but there are other considerations as well. The citric acid cycle is something that we describe as anaplerotic. A-n-a-p-l-e-r-o-t-i-c. Anaplerotic literally means to fill up. To fill up. What does that mean? It means that if we look at the cycle, we see several things. Citrate, we see alpha-ketoglutarate, we see succinyl CoA, we see oxaloacetate, all of which have links to other pathways. And we remember that these molecules, even though we like to think of them as only existing here, can interact with these other pathways very abundantly. What if I need to make a bunch of amino acids that are related to glutamate? What's the cell going to do? Well, it's gonna take some of this alpha-ketoglutarate; it's gonna swipe it out of the cycle; it's gonna make glutamate; and it's gonna make those other amino acids, ultimately, maybe purines, which are nucleotides, that the cell needs. What if the cell has just absorbed, okay, a big *** of protein or a big *** of nucleic acid? In that case, it's gonna say, "Well, I've got all this stuff over here. "Let's break it down, break it down, break it into glutamate, "and dump it in the cycle where it can be used." Now that turns out to be very important if we think about alternative diets. We don't technically have to eat glucose to have glucose. How might we get glucose from protein? Well, there's a good way. Let's say we started with a bunch of protein. We've got a bunch of glutamate. We dump it in the cycle. We make alpha-ketoglutarate. We make succinyl CoA. We make oxaloacetate. And look where we're going. Gluconeogenesis. We've just converted amino acids into glucose. If you're on a low glucose or a low carb diet and you're eating a lot of protein, you've got plenty of energy. Why? Because this is what you're doing. You can also get things out from this direction, okay. Coming in through aspartate; to oxaloacetate; and ***, up to glucose. Now what we can't do, and this would be really good for weight loss, what we can't do is we can't convert acetyl CoA into glucose in net amounts. We can't do that. "We" being higher animals. In fact, animals in general cannot convert acetyl CoA. Well, you say, "But look, if you run this all the way around, "why doesn't that work? "Why can't I make this into glucose?" Any thoughts? Yeah? Student: Because you need to attach the CoA to oxaloacetate to start the cycle in the first place. Professor: Okay. So he said you need to attach the acetyl CoA to the oxaloacetate. No, that's not quite right. What's happening in the cycle? We're adding two carbons. What's happening in the cycle? We're losing two carbons, right? To carbon dioxide. Do we have any carbons leftover to go and make more oxaloacetate? We start with one oxaloacetate; we end with one oxaloacetate, right? Do we have any net gain of oxaloacetate in this cycle? We don't. So even though we could take this one and make glucose, we're gonna run out of cycle pretty quickly if we just take it and we don't replenish it. So animals cannot make glucose in net amounts from acetyl CoA. And I say that's very important for dietary purposes. Why is that important for dietary purposes? It means I can't convert the fatty acids that I have in my body. I can't convert them, with rare exception, we'll talk about that later, but with rare exception, I can't convert them into glucose. If I could, I'd have a really good way of burning up an awful lot of fatty acids, which are components of fat, very quickly. I can't do that. It would be nice to be able to do that. To eat a fat diet and do what I'm doing with amino acids to make glucose, but I can't do it. It doesn't work. Everybody understand that? It doesn't work, because fatty acids break down to acetyl CoA. Fatty acids, in their breakdown, give acetyl CoA. So yeah, they're really great for burning in the citric acid cycle, but it sure would be nice to use those guys to make glucose as well. I can't do it. Now there are some organisms that can, in fact, do that. In order to do it, they have to do a trick. Or not a trick, they actually have to have a couple different enzymes than what we do. So I'm gonna show you those. Before I do that, are there any questions about the anaplerotic nature of the pathway? These arrows go back and forth. They feed in both directions. So we can put things in the pathway. We can take things away from the pathway. If we keep taking away from the pathway, though, we will eventually stop the pathway. Yes, sir? Male student: Question. Possibly enzymatic treatment of that, because like you say, you talked about people who are lactose intolerant ingesting the enzyme along with lactose so they don't have the digestion problems. Would that not be something that could be done with the introduced enzyme here because of its location within the cell? Kevin Ahern: You're talking about introduce the enzyme to use acetyl CoA to make glucose? Male student: Yeah, here is a diet shot. Professor: Yeah. So his question is could I introduce an enzyme into cells, in our cells, that would allow us to make glucose from acetyl CoA. The main problem with any of those is getting the enzyme in the cell. In the case of lactose intolerance, the enzyme is not crossing the cell barrier, but its in our intestinal fluid which is where the lactose is. So it doesn't have to cross the cell barrier. Anytime we try to cross the cell barrier with something, it's a totally different world. So no, the answer is we couldn't do that. If we wanted to do that, we would have to engineer cells to carry the coding for those enzymes, and I'm gonna show you those enzymes in a second, but we'd have to engineer the cells to have the coding so they could make that. You wouldn't want to mess with that. I can tell you that. The possible problems that you would have with that in terms of complexity because now these are enzymes that have never been in an animal cell before and how they would upset the metabolic pathways that are very carefully balanced, you probably wouldn't want to do. Female Student: Our bouncing friend. Kevin Ahern: What's that? Female student: Our bouncing friend. Kevin Ahern: I wouldn't call him a friend but you know, bouncing thing is back. Well, let's look at a couple of organisms, alright some organisms that use that. It's called the glyoxalate cycle. I know I'm jumping around a little bit. Try it again. "Kevin's gonna make us learn another cycle, and he's gonna do it before the period's over." Yeah, that's the bad news. The good news is you already know almost all of it. The reactions that you see here are the same enzymes that occur in the citric acid cycle except for two. Now, citrate synthase, we saw that in the citric acid cycle. In fact, these two cycles, the cycle I'm showing you here, the glyoxylate cycle, and the citric acid cycle both occur in plants, bacteria, and yeast. That's where these enzymes are present. So we see the citric acid cycle and we see the glyoxylate cycle both occurring. It's not one or the other, although we can think of situations where one or the other might be favored, and we'll talk about that in a minute. But this is occurring there, in the mitochondrion of these plants, yeast, and the cytoplasm of bateria. Now citrate synthase, same enzyme as citric acid cycle. Aconitase, same enzyme as citric acid cycle. Here's a new guy, alright. The new guy, called isocitrate lyase, you'll notice that isocitrate lyase pops up before the first oxidative decarboxylation can occur. In the citric acid cycle, we had isocitrate dehydrogenase present there. Here, we have isocitrate lyase. What's the difference? Well, isocitrate dehydrogenase catalyzed formation of carbon dioxide. We lost a carbon. This guy isn't doing that. It's catalyzing the splitting of this six carbon molecule into a four carbon molecule and a two carbon molecule. We haven't lost any carbons yet, and in fact, we're not going to. What happens to succinate? If I had the citric acid cycle going and I have the glyoxalate cycle going, what's gonna happen to succinate? The citric acid cycle, right? This guy's gonna go off, and it's gonna make oxaloacetate, right? How about this two carbon piece? What happens to it? Here's where the second enzyme comes in. The second enzyme is called malate synthase. It takes a second acetyl CoA, it combines it with glyoxalate, and makes a four carbon piece called malate. What happens to the malate? The malate goes to oxaloacetate. That's just the citric acid cycle again. Malate dehydrogenase. At this point, how many oxaloacetates do we have? We have two. How many carbons have we lost? Zero. So what we've done with the glyoxalate cycle, I shouldn't say we, because we aren't doing it. Plants, yeast, and bacteria are doing this. What they're doing with this is they are creating an extra oxaloacetate because basically they're taking an acetyl CoA and an acetyl CoA and ultimately converting it into an extra oxaloacetate. They're not losing the two carbons. What does this mean about these guys' ability to convert fat into glucose? They can do it, they can do it. Plants, yeast, and bacteria can convert acetyl CoA into glucose in net amounts because they get an extra one of these every time the cycle turns. Well, now we start thinking, "You said both cycles are going. "How do I know which one's going when?" Well, as long as we think of both of them going, we're in good shape. But might one be favored at one time versus another time? And the answer is, "Yeah, it probably is." When do you suppose this one will be favored compared to the citric acid cycle? Under what conditions would this guy work more than the other one would? Any thoughts? Yes, sir. Male student: Possibly at night. Kevin Ahern: Possibly at night if you're photosynthetic. The answer is yes, to some extent. But bacteria aren't all photosynthetic. There's a more general answer to that. Yes, Connie. Connie: Possibly high levels of acetyl CoA. Kevin Ahern: High levels of acetyl CoA will contribute to it. That's true, too. There's a better answer yet. Yes, Joseph. Joseph: To get glucose. Kevin Ahern: What's that? Joseph: To get glucose. Kevin Ahern: That will contribute to it, too. That's still not what I'm after. Male student: Winter. Kevin Ahern: Winter, maybe. I would say probably not winter, no. Female student: How much energy there is. Kevin Ahern: She says, "How much energy there is." What does that mean? Female Student: Like when there's high energy, the citric acid cycle will go. When there's low energy, it won't. What was the other factor in controlling the citric acid cycle? You're saying energy, and I used... Female Student: ATP. Kevin Ahern: Well, I'm not saying even ATP. Female Student: ***. Kevin Ahern: ***, right? Which cycle's gonna need more ***? Kevin Ahern: The citric acid cycle. So the one that's the most dependent upon *** is not gonna be running when there's plenty of NADH, right? This will tend to run when there's less ***. That makes really good sense, guys. Think about pathway integration here. If there's a lot of NADH... Let's see, what did we need to make glucose? In gluconeogenesis we needed triphosphates, we needed pyruvate, and we needed NADH. Now we can use that NADH to go and synthesize glucose. We've got a way of reducing our NADH concentration by making this and then going back up to glucose. That's pretty cool. There's balance that is happening in these cells based on need. Now, questions about that? Yes? Male student: [Inaudible] Kevin Ahern: What is... So his question is, "What's different about animals? "Why don't animals have this cycle?" Animals don't have these two enzymes, malate synthase and isocitrate lyase. These two enzymes are missing in animals. They don't have a way of using acetyl CoA without losing those two carbons every time the cycle turns. This is bypassing those loss of two carbons, and that's why we're ending up with an extra oxaloacetate. Male student: [Inaudible] Kevin Ahern: His question is, "Why don't we have this?" And the answer is because it doesn't work for us. That's not a very satisfying answer, but it's true. That's basically what it is. If there were an evolutionary advantage for us having this, we would have it. Animals have very different energy needs than do plants, yeast, and bacteria. Remember we talked about glycogen metabolism and how we needed that quick energy? They don't need that. So they have evolved very different kinds of situations than what we had. I know that is not a very satisfying answer, but unfortunately, that is the answer to your question. Do I see a hand? Male student: [Inaudible] Kevin Ahern: What? Male student: [Inaudible] So if we talk about seafloor plants, what you're talking about, are seafloor plants different as a result of this? And seafloor plants actually have a slightly different scheme but this doesn't affect it tremendously. No, okay. This is a phenomenon true in plants in general. Seafloor plants are using a different way of getting carbon dioxide in, but that's the main thing they're doing with that. So that doesn't affect this in any significant way. Male student: Does fungi have two different cycles, too? Professor Ahern: Do fungi... Yeah, fungi have two different...they have the glyoxalate cycle as well. So I say yeast. I use that just as fungi in general. Fungi, bacteria, and plants. So that's cool. Now I skipped over a couple things. Let me finish up those couple things, and we'll call it a day. So one of the things I didn't talk about was pyruvate dehydrogenase complex regulation. I'm gonna keep it simple, as I said. Let's talk about it in simple terms. Oh boy, there's our friend phosphorylation again. This enzyme... Why, I seem to have the worst luck with laser pointers. It said that it was working, but it's not working. This enzyme, pyruvate dehydrogenase, is regulated partly by phosphorylation. In the unphosphorylated form, it's the most active. In the phosphorylated form, it's the least active. Not surprisingly, there's a kinase that converts between the two. Not surprisingly, there's a phosphatase that takes the phosphate off. This enzyme is also regulated partly allosterically. And I hate this figure, by the way. It makes it look way more complicated than it is. Basically we'll be concerned right here with this guy, ATP. ATP turns this enzyme off. Allosterically turns that enzyme off. Everything else on there, you can ignore. And again, high energy, do we want to be oxidizing our pyruvate and burning energy? No, we don't. We want to have that enzyme turned off. This is a very good way of controlling the entry of pyruvate and acetyl CoA into the citric acid cycle. Okay, the last thing I'll talk about is right here. Arsenic, arsenic poisoning. Everybody knows arsenic is really nasty stuff. Arsenic will kill you. There's a couple ways arsenic can kill you, and this is one of them. Arsenic can readily be converted in our bodies into arsenite, which is what you see on the screen there. And arsenite has the very unfortunate chemical property that it will react with the sulfhydryls of lipoic acid or lipoamide. When it does that, where was lipoamide important again? Two enzymes. Pyruvate dehydrogenase and alpha ketoglutarate dehydrogenase. The two that have the same mechanism, right? Those are pretty important. One gives us carbons for the citric acid cycle. The other is for the citric acid cycle itself. If we stop the citric acid cycle, what's gonna happen to our energy metabolism? We're dead, okay. So we get the formation of this destroyed lipoamide. Fortunately, if we catch it early enough, one of the ways we can treat it is with this compound called BAL. And that's what I'm gonna call it because I never remember the full name of it anyway. BAL will basically displace the arsenite from the lipoamide and result in a regenerated lipoamide. So BAL is basically taking it away, as we can see here, taking it away from the lipoic acid and leaving us back with lipoic acid so that we can do things. There are other problems that arsenic causes us. This is not the only one, but this is one way that we can treat arsenic poisoning. Yes, sir. Male student: Do you know of any bacteria they found in Mono Lake that are very arsenic tolerant? Male student: Is something like this native in them? He's got a very good question. About a year ago there was a provocative paper that was published in I believe Nature that claimed that there were bacteria in Mono Lake in California that were using arsenate as a replacement for phosphate in making nucleic acids. That has largely, it's not completely proven, that largely has not stood up to scrutiny at this point. And so they're still actively investigating these unusual bacteria, but it appears there's not solid evidence yet that that's happening. So that's as much as I can tell you at this point. Alright, we're gonna finish early again. Have fun and see you on Friday. [classroom chatter] [END]