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Biochemistry 4 Biology > Introduction to Biology
Video Lectures - Lecture 5/nTopics covered: /nBiochemistry 4/nInstructor: /nProf. Robert A. Weinberg/nTranscript - Lecture 5/nGood morning, class./nI just wanted to spend the first couple minutes clearing up three issues. None is a major conceptual issue, but we like to focus on details and get them right, get them correct here as well. Firstly, I misdrew a reaction last time that described why RNA is alkali labile, i.e., if we have high pH we call that an alkali pH, or an alkaline pH, actually, to use the adjective./nAnd we said that hydroxyl groups can cause the cleavage of the phosphodiester bonds of RNA but not DNA. And the way I described that happening is that the alkali group causes the formation of this five-membered ring right here, two carbons, two oxygens and a phosphate. And that resolves eventually to this where there's no longer any connection with the ribonucleoside monophosphate below./nAnd I drew it like this, without an oxygen, and that's a no-no because, in fact, in truth, and as many of you picked up, this reverts to a two prime hydroxyl. So, please note there's a mistake there. There's also a couple other mistakes. For example, in the textbook it gives you the impression that when you polymerize nucleic acids you use a monophosphate to do so./nAnd, if you listened to my lecture last time, that doesn't make any sense, because you need to invest the energy of a triphosphate in order to create enough energy to generate enough energy for the polymerization. The textbook is incorrect there. Textbooks are written by people, for better or worse, and as such, like everything else, they are a mortal and fallible. So, the truth of the matter is, when you're polymerizing DNA or RNA you need one of the four ribonucleoside or deoxyribonucleoside triphosphates in order to donate the energy that makes possible this polymerization./nAnd please note that is a mistake in the book. Recall, as I said last time, the fact that ATP is really the currency of energy in the cell, and that its energy is stored and coiled up in this pent up spring where the mutual electrostatic repulsion between the three negatively charged phosphates carries with it enormous potential energy./nAnd some of that potential energy can be realized, during the synthesis of polymerization of nucleic acids by cleaving this bond here. One can also generate potential energy by cleaving this bond here. This is the alpha, the beta and the gamma-phosphate. And cleavage of either can create substantial energy, which in turn can, as we'll indicate shortly, be invested in other reactions. The reaction of polymerization./nA second point I'd like to make to you is the following, and you'd say it's kind of coincidence. The currency of energy in the cell is ATP, adenosine triphosphate, we see its structure here, and this happens to be one of the four precursors of the RNA. So, the same molecule is used in these two different ostensively unrelated applications. One, to polymerize to make RNA where genetic information is stored and conveyed./nOr, alternatively it's used here in this context in order to serve as a currency for energy. High energy as ATP. ADP with a little lower energy. AMP monophosphate with even lower energy. And you might ask yourself, scratch your head and say why is the same molecule used for these two different things? In fact, there are yet other applications of these ribonucleosides which also seem to be unrelated to the storage or the conveyance of genetic information./nAnd it is believed, probably correctly, that the reason why the same molecule is used for these totally different applications is that early in the evolution of life on this planet there really were a rather small number of biological molecules that existed. Indeed, as we'll mention again later, it's probably the case that the first organisms didn't use DNA as genomes. It's an article of faith with us that one stores genetic information in DNA molecules./nAnd I implied that quite explicitly last time. But, the fact of the matter is, it's probably the case that the first organism, the first pre-cellular life forms used RNA as the genetic material, RNA to store things, replicating RNA via double-stranded RNA molecules as a way of archiving genetic information. And only later during the evolution of life on this planet, when that later was we can't tell, but it could have been a hundred or two hundred years later./nObviously, if we're talking about the origin of life as between 3.3 and 3.5 billion years ago, we can't really localize that in time very well, but only later was DNA assigned the job of storing, in a stable fashion, genetic information. And as a consequence, we come to realize as well yet another discovery, which is that all the catalysts that we're going to talk about today, the enzymes as we call them, almost all modern-day enzymes are proteins./nAnd we talked about them briefly before. But over the last 15 years, 20 years there's been the discovery that certain RNA molecules also posses the ability to catalyze certain kinds of reactions. When I was taking biochemistry, if somebody would have told me that, I would have called the psychiatric ward because that was such an outlandish idea. How can an RNA molecule catalyze a biochemical reaction?/nIt doesn't have all the side groups that one needs to create the catalytic sites for reactions. But we now realize, on the basis of research which actually led to a Nobel Prize being awarded about five years ago, that RNA molecules are able to catalyze certain kinds of reactions. And that begins to give us an insight into how life originated on this planet because RNA molecules may have stored genetic information, as I said before, RNA molecules, or their precursors like ATP, may have been their currency for storing high energy bonds, as is indicated here./nAnd RNA molecules may well have been the first enzymes to catalyze many of the reactions in the most primitive life forms that first existed on this planet. And, therefore, what I'm saying is that as life developed in the first hundred or two hundred million years, who knows how long it took, gradually DNA took over the job of storing information from RNA and gradually proteins took over the job of mediating catalysis, of acting as enzymes to taking over the job from RNA molecules./nToday there are certain vestigial biochemical reactions which we believe are relics, echoes of the beginning of life on earth, which are still mediated by RNA catalysts. We think that they are throwbacks to these very early steps, maybe even in pre-cellular life form where RNA was delegated with the task of acting as a catalyst./nWe're going to focus a lot today on the whole issue of biochemical reactions and the issue of energy. And this gets us into the realization that there really are two kinds of biochemical reactions. Some of you may have learned this a long time ago. Either exergonic reactions that release energy, that produce energy as they proceed, or conversely endergonic reactions which require an investment of energy in order to move forward./nSo, here, obviously, if this is a high energy state and we're talking about the free energy of the system, which is one way to depict in thermodynamic language how much energy is in a molecule, if we go from a high energy state to a low energy state then we can draw this like this and we can realize that in order to conserve energy, the energy that was inherent in this molecule, the high potential energy is released as this ball or this molecule rolls down the hill./nAnd, therefore, the reaction yields energy, it's exergonic. And, conversely, if we want this reaction to proceed, we need to invest free energy in order to make it happen. The free energy happens to be, more often than not, in the form of chemical bonds, i.e., energy that can be invested, for example, by taking advantage of the potential energy stored in these phosphodiester, in these phosphate-phosphate linkages indicated right here./nHere, by the way, is the space-filling model of ATP just for your information. That's the way it actually would look in life, and this is the way we actually draw it. Now, having said that, if we look at the free energy profile of various biochemical changes then we can depict them, once again, in this very schematic way here./nAnd, by the way, free energy is called G, the Gibbs free energy after Josiah Gibbs who was a thermodynamic wiz in the 19th century at Yale in New Haven. And here what we see is that the change in free energy between the reactants and the products is given by delta G. So, by definition, we start out the reaction with reactants./nAnd we end up at the end of the reaction with products. And, overall, if the reaction is exergonic and will proceed forward, it releases energy. And the net release of energy is indicated here by delta G. But, more often than not, biochemical reactions that are energetically favored, that are exergonic actually can't happen spontaneously. They don't happen spontaneously because, for various reasons, they have to pass through an intermediate state./nWhich actually represents a much higher free energy than the initial reactants posses. And this higher free energy, that they need to acquire in order to move over the hill and down into the valley, is called the energy of activation, the activation energy. And, therefore, if I were to supply these reactants with energy, for instance, let's say I were to heat up these reactants and therefore give them a higher degree of thermal energy which they might be able to use to move up to this high energy state./nI supplied them with free energy by giving them heat. Then they might be able to move up to here and then roll down the hill. But in the absence of actually actively intervening and supplying them that energy, they'll remain right here, and they may remain right there for a million years, even though in principle, if they were to reach down here, they would be much happier in terms of reaching a much lower energy state./nTo state the obvious, all these kinds of reactions wish to reach the lowest energy state possible. But in real-time it can't happen if there is a high energy of activation. Now, what do enzymes do? As always, I'm glad I asked that question. What they do is they lower the energy of activation. And this is in one sense obvious, and in one sense it's subtle, because enzymes have no affect on the free energy state of the reactants, they have no affect on the free energy of the products./nAll they do is to lower the hump, and they may lower it very substantially. And because they lower it substantially, it might be that some of the reactants here may, just through a chance, acquisition of thermal energy, be able to move over the much lowered hump and move down into this state right here. Now, the actual difference in the Gibbs free energy is totally unaffected./nAll that happens is that the enzyme, by lowering the energy of activation, make this possible in real-time. The fact is that ultimately, if one were to plot many kinds of reactions, many reactions, as is indicated here, have a very high activation energy, and therefore we look at it like this. But there could be other reactions which might have an activation energy that looks like this, almost nothing at all./nAnd these reactions could happen spontaneously at room temperature in the absence of any intervention by an enzyme. For example, let's say we're talking about a carboxyl group which discharges a proton. We've talked about that already. Well, that reaction happens spontaneously at room temperature. It doesn‘t need an enzyme to make it happen. It can happen because there's essentially not energy of activation./nBut the great majority of biochemical reactions do have such an activation energy, and therefore do require a lowering like this in order to take place. Now, let's imagine other versions of the energy profile of a reaction. And keep in mind that what I'm showing here on the abscissa is just the course of the reaction. You could imagine I'm not really plotting time./nI'm just talking about a situation where to the left the reaction hasn't happened and to the right it has happened. Can you see this over there? Then I won't write over there. All right. Let's see if this works./nBoy, here we are in the 21st century and we still haven't worked this out. OK. Everybody can see this right here, right? OK. So, look. Let's imagine we have a reaction that looks like this, a reaction profile that looks like this, where these two energies are actually equivalent. OK? I've tried to draw them on./nWell, they're not exactly, but they're pretty much on exactly the same level. And let's say we start out with a large number of molecules right over here. Now, if there were an enzyme around, the enzyme might lower the activation energy and, in so doing, make it possible for molecules to tunnel through this hill and move over here. The fact that when a molecule gets over here it has the same free energy as over there means that the catalyst may, in principle, also facilitate a back reaction./nWhat do I mean by a back reaction? I mean going in exactly the opposite direction. And so, once molecules over here are formed, the energy lowering affects of the enzyme may allow them to move in both directions. And, therefore, what we will have is ultimately the establishment of an equilibrium. If these two energy states are equivalent then, I will tell you, 50% of the molecules end up here and 50% of the molecules end up here./nAnd here we're beginning now to wrestle between two different independent concepts, the rate of the reaction and the equilibrium state of the reaction. Note that the enzyme has no affect whatsoever on the equilibrium state. These two are at equal free energies, the equilibrium state. Whether the energy barrier is this high or whether it's this high is irrelevant./nThe fact is if the enzyme makes possible this motion back and forth, the ultimate equilibrium state will be 50% of the molecules here and 50% of the molecules there. And, therefore, the enzyme really only affects the rate at which the reaction takes place. Will it happen in a microsecond or will it happen in a day or will it happen in a million years? The enzyme has no affect whatsoever on the ultimate end product, which in this case is the equilibrium./nOf course, there is a simple mathematic formalism which relates the difference in free energies with the equilibrium. Here we might have a situation where 80% of the molecules end up at equilibrium over here and 20% end up here. Or, we might end up as a state where 99.9% of the molecules end up here and 0.1% of the molecules end up here./nBut that ultimate equilibrium state is no way influenced by the enzyme. They just make it happen in real-time. And, therefore, to repeat and echo a point I made last time, if most biochemical reactions are to occur in real-time, i.e., in the order of seconds or minutes, an enzyme has to be around to make sure they happen. In the absence of such an enzyme of its intermediation, it just won't happen in real-time./nEven though, in principle, it's energetically favored. So, let's just keep that very much in mind in the course of discussions that happen. And let's just begin now to look at an important energy-generating reaction in the cell which is called glycolysis. We already know the prefix glycol. Glyco refers to sugar. And lysis, L-Y-S-I-S refers to the breakdown of a certain compound. I am not going to ask you, nor is anyone else in the room going to ask you to memorize this sequence of reactions./nBut I'd like you to look at it and see what take-home lessons we can distill out of that, what wisdom we can learn from looking at such a complex series of reactions. Perhaps, the first thing we can learn is that when we think about biochemical reactions, we don't think of them as happening in isolation. Here I'm talking about, for example, in this case I could be talking A plus B going to C plus D, and there might be a back reaction to reach equilibrium./nAnd we're just isolating that simple reaction from all others around it. But in the real world in living cells most reactions are parts of very long pathways where each of these steps here indicates one of the others, a step in the pathway. What we're interested in here is how glucose, which I advertised two lectures ago as being an important energy source, is actually broken down./nHow does the cell harvest the energy, which is inherent in glucose, in order to generate, among other things, ATP, which we've said repeatedly is the energy currency? ATP is used by hundreds of different biochemical reactions in order to make them happen. These other biochemical reactions are endergonic, they require the investment of energy, and almost invariably, but not invariably, but almost invariably the cell will grab hold of an ATP molecule, break it down usually to AMP or ADP./nAnd then utilize the energy, which derives from breaking down ATP, it will invest that energy in an endergonic reaction, which in the otherwise would not happen. So, here we reach the idea that perhaps by investing energy in a reaction, the equilibrium is shifted. Because by investing energy, actually, the cell is able to lower the free energy state between these two./nAnd that makes it possible for their equilibrium to be much more favored. Let's look at this glycolytic pathway. Glycolytic refers, obviously, to glycolysis. And here we start out with glucose. We're drawing it out flat rather than the circular structure we talked about last time. And let's look at what happens here, again, not because anyone wants you to memorize this, but because some of the details are in themselves very illustrative./nThe goal of this exercise is to create ATP for the cell, but the first step in the reaction is actually totally counterproductive. Look at the first thing that happens. The first thing that happens is that the cell invests an ATP molecule to make glucose-6-phosphate. I've advertised the goal of this is to generate ATP from ADP, adenosine diphosphate. But the first thing here, this is an endergonic reaction in which the cell invests energy to create this molecule here./nSo, this doesn't make sense. But ostensively it must make sense, at one level or another, because you and I, we're all here, and everybody in this room, at least this moment is metabolically active. All right. So, we've got this molecule here, glucose-6-phosphate. And this can isomerize. You see, here's glucose-6-phosphate, fructose-6-phosphate. And, the fact of the matter is, there's no oxidation reduction reaction here./nIt's just an isomerization. And this molecule and this molecule are virtually in the same free energy state. It happens to be the case that their profile will look very much like the one I drew you before. Their energy profile will look like this. And one needs an enzyme to lower it, but there's no energy that needs to be invested in converting one to the other because they're very similar molecules and therefore incomparable free energy states./nNow look at the next step. The next step is again another ostensively totally counterproductive way of generating energy. Because, once again, ATP, the gamma-phosphate, its energy is invested in creating a dephosphorylated hexose, fructose 1, 6-diphosphate where the numbers refer obviously to the identities of the carbon. And now we have a dephosphorylated fructose molecule./nAnd so here you can actually see what the three-dimensional, what we would imagine closer to what the three-dimensional structures of these molecules look like. And we shouldn't focus this time on whether it's this or this. For all practical purposes, let's just focus on this pathway here. And here, for the first time, what now happens is that this hexose is broken down into two trioses, i.e., into two three carbon sugars. And this is a slightly exergonic reaction./nIt yields, it happens without the investment of energy. And there's an enzyme, once again, that's required in order to catalyze it. But let's be really clear now. Now we have to follow the fate of two molecules. The first triose and the second triose. They have different names, but we're not going to focus on the names. One thing you notice about these trioses is that they're readily interconvertible. Once again, we can image that we have a situation that looks like this./nThese are flipping back and forth. And therefore, for all practical purposes from our point of view, these two are equivalent because they can be exchanged virtually instantaneously one with the other. Now, so far we've actually expended energy. We haven't harvested energy. But, keep in mind, the old economic dictum; you have to invest money to make money. And that's what's going on here. The first thing that happens is we have an oxidation reaction./nWhat's an oxidation reaction? We want to strip some electrons, a pair of electrons off of this particular triose, the 3 carbon sugar. And by stripping off a pair of electrons we donate the electrons from NAD+ to NADH. And here these structures are given in your book. But NADH, it turns out, is the electrons are pulled away from the triose and they're used to reduce NAD+ to NADH./nKeep in mind that in an oxidation reaction, one molecule that's being oxidized is deprived, is denied a pair of electrons. The other molecule that's being reduced, in this case NAD, acquires a pair of electrons. And you can focus, if you want, about the charge of these molecules, one or the other. But, keep in mind, that in these oxidation reduction reactions, whether it's plus charged or minus charged is irrelevant. The real name of the game is the electrons./nForget about the protons, whether it has a plus charge or it's neutral. The real name of the game here is that two electrons are being used to reduce this molecule to this. By the way, third mistake I forgot to tell you before, there's a double-bond in one of the pyrimidines in the book that doesn't make any sense. Whoever finds it gets a prize, but no one's figured out what the prize is yet. So, this double bond gets reduced. You see the difference between this and this over here. And this NADH, it turns out, is a high energy molecule./nThe street value of NADH is three ATPs, i.e., in the mitochondria NADH can be used to generate three ATPs, and that's worth something. So, NADH on its own is a high energy molecule. It can't be used for that many things, but it can be pulled into the mitochondria where it's converted to three ATPs. So, we say, well, we're starting to make some money out of this investment because we've made, in fact, these NADHs./nSee right here. Why do we say two NADHs? Because there are two trioses we're working with, and each one of the trioses gives you an NADH. So, everything that's going on after this, starting from the top here, is now double because we're looking at the parallel behaviors of two identical three carbon sugars. So, here we've so far generated, in principle, six ATPs./nHow much did we invest already up to this point? Two. We invested two but we harvested six. Already we're starting to make a little money because I told you the street value of an NADH is three ATPs on the black market. OK, so what happens next? Next is another good thing. Each of the trioses, one can actually cause each of the trioses to generate an ATP molecule from an ADP./nWhat happens here? It turns out that this phosphate over here is actually in a pretty high energy state, in no small part because of electron negative-negative repulsion. And by stripping this phosphate off this high energy phosphate stripped off of this molecule here, whose name we will ignore, allows us to phosphorylate an ATP. And since there are two trioses being converted, we're going to get two ATPs./nSo, in effect, now we're actually ahead. We started out investing two, we got six back from the NADHs, and we're getting two back here. So, we've made two ATPs. This is a good thing. Keep in mind, ADP is lower energy, ATP is a high energy. Once again, we have an isomerization where these two molecules are at comparable states here and here, where the phosphate just jumps over to this state./nAnd this hydrolyzes spontaneously and we get this molecule right over here, phosphoenolpyruvate at the end. And, once again, we harvest two ATPs, one ATP from each of the trioses. And we end up, at the end of this reaction, with pyruvate. And you'll say this is terrific because we invested two ATPs, we harvested four, plus we got six from the NADHs, right?/nTwo NADHs, each NADH gives us three each, so let's do the arithmetic. Let's do the balance sheet. We invested to begin with, with the one glucose, we invested two ATPs. That was early on. Then the return was first two NADHs, which I've told you equals six ATPs./nBecause an NADH is worth three ATPs. This is so far good. And now subsequently we've made four ATPs so that the net yield looks pretty useful. Six plus four is ten minus two, a profit of eight ATPs from one glucose molecule. This is terrific you may say, but there's a rub./nThere's a catch. If glycolysis is occurring in the absence of oxygen, if that happens, then we have a problem here, because the only way that these NADHs can generate ATP is if there is oxygen around to take these electron pairs and use them to reduce an oxygen molecule. That is, by the way, part of the reason we breathe./nKeep in mind that when you generate an NADH from an NAD molecule, you need to regenerate the NAD. You can't just accumulate more and more NADHs. You need to regenerate the NAD. And, therefore, this NADH, with their electron pairs, the electron pairs have some to be disposed of. You have to regenerate NAD. You can't just make more and more and more of this. So, how do cells get rid of it? Well, how they get rid of it is simple./nYou take the electron pairs and you slap them onto oxygen, and that's really called combustion. And you get a lot of energy out of that. But what happens if all of this is occurring anaerobically? Anaerobically means the reaction is occurring in the absence of oxygen. Well, if you have a yeast that's growing 14 feet underground, this is happening anaerobically./nIf you have a yeast that's fermenting in a big keg to make wine or beer, it's also probably happening anaerobically. If you start running in a 100 yard sprint, or let's say you had to run a mile, then initially there's enough oxygen, there's a lot of oxygen around to allow you to get rid of these NADHs and dump the electrons that they have acquired onto the oxygen molecule. And that's fine. That's worth a lot because, in effect, what you're doing is you're taking oxygen and hydrogen and you're combusting them together./nAnd that's great. But as you start running down the street, soon the oxygen supply to your muscles is going to run out, and soon a lot of the energy production in your muscles happens anaerobically. Why? Because you can't get oxygen quickly enough to your muscles, and therefore, for a period of time, you start feeling that burning sensation in your muscles because oxidation of NADH isn't happening./nAnd these NADHs instead are regenerated by another way. How are they regenerated? The electron pairs of the NADHs, must be, are dumped back onto this molecule right here, pyruvate. They're not used to make ATP because they can't be used to make ATP because there's no oxygen around to accept the electron pairs that these NADHs have acquired./nAnd so, what happens with these valuable NADHs? Under anaerobic conditions this doesn't happen. These NADHs are used instead, their electrons are donated to our friend pyruvate here, these three carbon sugar. And what happens, when they are donated back to the pyruvate, in order to regenerate NAD you need more NAD to pick up to use later in the reaction, to use over again in another reaction./nWhen you donate the electrons from NADH back onto pyruvate, what happens? You get lactic acid. Lactic acid is what makes your muscles burn when you're running very quickly and you can't get enough oxygen into them to begin to burn up the NADH. So, instead of using NADH to generate ATP, it's diverted to make lactic acid. That's in one sense good because you regenerate NAD./nWhy do you need to regenerate NAD? Because you need a lot of NAD around for the earlier steps in the reaction. Keep in mind, early in the reaction you need NAD here. If you don't regenerate it then glycolysis grinds to a halt. So, even though you make NADH and it's a good thing in principle, in practice it has to be recycled. And if it's not recycled to make more new NAD to allow this step to happen then the whole glycolytic reaction will shut down and you're in a mess./nHowever, sadly, in the absence of oxygen, the only way to recycle this is to dump these electrons not onto oxygen which is energy rich, it's dump them back onto pyruvic acid creating lactic acid. So, you reduce this bond right here. So, you get CH, COH. This bond right here is reduced and you get lactic acid./nSo, instead of a carbonyl bond here you have CH and COH right here, that's a reduction reaction. And now you're able to regenerate the NAD. And now you say that's a great thing. But, keep in mind, that now the entire glycolytic reaction, how much is our net profit now? Before I was gloating about the fact that we made eight ATPs, we netted eight ATPs out of this. What are we back down to now? What's the whole net yield now?/nWell, the TAs can't answer. It's two, because we invested two and we got out four. And it's only two. Now, why is this so interesting? Well, until about six hundred million years ago there wasn't that much oxygen in the atmosphere. And in the absence of oxygen this is almost the only reaction that could be used in order to generate energy. And about six hundred million years ago more and more oxygen from photosynthesis became dumped into the atmosphere./nAnd soon oxygen became available to organisms like our ancestors. And then they could actually begin to recycle this NADH in a much more productive way. And as a consequence what happened, instead of having glycolysis yielding two, we could go up to this theoretical eight because the NADHs could now deposit their electrons on oxygen, which is much more profitable./nIn fact, I've just told you now that in the absence of oxygen you can only make two ATPs. I will tell you, without providing it to you, that in the presence of oxygen you can make 34 ATPs. And 34 is, we can agree, much better than two in the presence of oxygen./nHigher life forms could not evolve until this much more effective way of generating energy became available. And, therefore, if our ancestors who lived longer than six hundred million years ago were very sluggish and they weren't very smart, the reason why they were sluggish and they weren't very smart is because they couldn't generate the energy that was required to efficiently drive metabolism. The metabolism, anaerobic metabolism, i.e., occurring in the absence of energy, is extremely inefficient./nIt just doesn't happen very well. Now, what actually happens if we have oxygen around? Well, what happens is something like this. We take the pyruvate, which is the product of glycolysis and which is this much more primitive pathway, and we dump it into the mitochondria./nAnd now we generate through this cycle here, which I'm not asking you memorize, please, don't do that. We generate the reactions which go from here and get us up to this 34 ATP yield per glucose. And the essence of the citric acid cycle, which happens in the mitochondria, keep in mind that the mitochondria look like this./nKeep in mind that the mitochondrion are the decedents of bacteria which parasitized the cytoplasm of cells probably 1.5 billion years ago. But if we now look at what happens in the mitochondrion, the pyruvate that we generated in the cytosol, in the soluble part of the cytoplasm is now pumped into the mitochondria, and there's a whole series of reactions that go on here, which takes this three-carbon sugar./nThe first thing that happens is that carbon is boiled off. Carbon dioxide, that's released. Now we're down to a two carbon sugar. And then this two carbon sugar is added to a four carbon sugar and progressively oxidized. And as it's oxidized what's spun off? Well, what's spun off is, for example, there's NADH which is spun off, there's ATP. See, there's an NADH which is spun off. Here's an NADH that's spun off. Here is a cousin of NADH./nIt's called FADH which, once again, generates a high energy molecule. Once again, the carbon molecules are oxidized, electrons are stripped away and used to create these high energy molecules, FADH and NADH. By the way, FADH, a cousin of NADH, is only worth two ATPs on the open market. Whereas, NADH, as I've told you repeatedly, is worth three. And by the time we add up all of the NADHs that have been generated by this cycling and the carbon dioxides that are releases, at the end of this cycle here we start with two carbons, add it to four and we get a six carbon molecule./nWe spew off some carbon dioxides here and go back to four carbon sugar. Add another two, go up to six carbons. Go around again, spin around the wheel. And each time we do that we generate a lot of NADHs, we generate a lot of FADHs, and we generate a lot of ATP./nIn all cases, these are highly profitable reactions simply because the NADHs and the FADHs can be used in the mitochondrion to generate ATP. So, let's look at the energy profile of the entire thing. Put it all together. This is where we started out at the beginning, and this is the end of glycolysis, OK?/nSo, now we're adding up the energy profiles of the whole sequence of reactions that constituted glycolysis, which begins up here and ends right here because pyruvate, as you will recall, is the product of glycolysis, the first step. The Krebs Cycle happens, or sometimes it's called the Citric Acid Cycle. So, let's just get these words straight. Citric Acid Cycle because it happens to be one of the cycles, or it's sometimes called the Krebs Cycle after the person who really discovered it, Krebs./nThe Krebs Cycle begins here. You see how the shading changes from pyruvate. And here we go all the way down there. And let's now look at what happens in terms of energy exchange. Recall that early on we needed to invest ATPs to kick up the energy state up to here. We invested ATPs at this stage right here, and then we began to get some back./nWe got these two NADHs, one NADH coming from each of the three carbon sugars. We got some more ATPs here and we got some more ATPs here, but these NADHs could not be used productively for generating ATP in the absence of oxygen, but in the presence of oxygen now we can begin to use these very profitably. Each of these makes three ATPs and each of these, obviously, makes ATPs./nAnd then let's look at what happens in the mitochondrion. Keep in mind here's the borderline between the cytosol, the cytoplasm and the mitochondrion. Here is where the oxygen is actually used and here we generate all these NADHs here, here and here, FADHs. And I keep saying, and it's still true, just in spite of the fact I keep saying it, that these NADHs can be converted to ATPs, and the ATPs can then be diffused, transmitted throughout the entire cell where they're then used invested in endergonic reactions./nHere we see all these NADHs. And look at the overall change in free energy. The initial steps in glycolysis didn't really take advantage. Glucose has inherent in it almost 680 kilocalories per mole of energy. It's pretty high up here. But by the time we get from here down to here, there's an enormous release of energy, it's harvested in the form of these molecules which are then reinvested./nIn the absence of oxygen, this entire procedure can only go from here down to here. And a lot of this drop from six to seven is futile because we have to reinvest this NADH. These cannot be used, actually, to generate more ATPs, as I've said repeatedly. So, this means in the end that we can generate an enormous amount of energy in the form of these coupled reactions./nHaving said that, let's actually look at what happens inside of the mitochondria. Inside of the mitochondria there are actually different physical compartments. See the blue space there, the intermembrane space, the blue spaces there? The matrix is on the inside./nThe intermembrane space is between the two, the inner and the outer membrane, and outside is the cytoplasm. The outer membrane, the inner membrane, in between it. So, look what happens, actually, in the mitochondrion. Those NADHs are used to pump protons from the inner space of the mitochondrion into the intermembrane space. I'm not showing you that happening./nBut you'll have to take it on my word. So, protons pictured here are extracted from NADH and FADH, and they're used to pump protons out here. And, therefore, protons are moved from here to here. Obviously, when you pump protons out the pH gets lower on the outside than it does on the inside, and because there's a gradient, there's a higher concentration of protons here than on the inside./nThe protons begin to accumulate outside here in the intermembrane space. Are they in the cytoplasm? No. They're in the space between the inner and the outer membrane. You start to accumulate in this blue space lots of protons. And this pumping of protons into the space between the two membranes requires energy, and the energy comes from our friends NADH and FADH as it turns out./nThey are responsible for causing this accumulation of protons in the space between the inner and the outer membrane. So, now we get lots of protons out there. And what happens now, the protons like to flow back in because there is a higher concentration here as they are inside the space that's called the mitochondrial matrix, on the inside of the mitochondrion. So, what happens?/nHere, yet another Nobel Prize winning discovery is the discovery of a very interesting molecule, or complex of proteins I should say, that looks in three-dimensions roughly like this. And what this complex does is as the protons flow through the inner channel here, they're moving down an energy gradient. They're going from a state of high concentration to a state of low concentration. What that does, that diffusional pressure actually yields energy./nAnd this complex right here harvests that energy in order to convert ADP into ATP. So, when I talk about NADH as being worth, each of them being worth three ATPs, what I'm really talking about is the fact that NADHs can be used to pump protons in the mitochondria outside here, and these protons can then be used, can then be pumped, can then flow in this way through this proton pump, which then uses ADP in the inner cavity of the mitochondria to create ATP./nAnd here we get finally the conversion of ADP into ATP. We can realize, finally, this much promised benefit. And then these ATP molecules are exported from the mitochondria throughout the entire cell and used to drive many reactions. We've already encountered one important set of reactions, and those reactions are the polymerization of nucleic acids./nNow, one final point I want to make is the following. We've just talked about metabolic, we've talked about the pathway of energy production in the cell. And you might have had the illusion, for a brief instant, that those are all, that's the sum of all the biochemical reactions in the cell. But, in fact, if we plot out all the biochemical reactions in the cell, they're much more complicated. Here is the glycolytic pathway./nYou see it right down here where nothing is named? Here is the Krebs Cycle right here. And we're not even talking about energy here. And as molecules move down this pathway from here to here to here to here, some of these molecules are diverted for other applications. Not for energy production but for other applications. And what happens out here, they are converted through a series of complex biochemical steps into other essential biological molecules./nWhat do I mean by that? If you give E. coli, a bacterium, you give it a simple carbon source like glucose and you give it phosphate and you give it a simple nitrogen source like ammonium acetate or something, E. coli can, from those simple atoms generate all the amino acids, can generate the purines and the pyrimidines, can generate all kinds of different complex biological molecules just from those simple building blocks./nAnd so, the process of biosynthesis involves not only the creation of macromolecules, these steps of what are called intermediary metabolism are used to synthesize all the other biochemical entities that one needs to make a cell. They're used to synthesize purines and pyrimidines./nThey're used to synthesize lipids, they're used to synthesize amino acids, and they're used to synthesize literally hundreds of other compounds. And when we see this chart like this, and nobody on the face of the planet has ever memorized this chart, each one of these steps, going from one molecule to the next, represents another biochemical reaction. And the vast majority of these biochemical reactions going from A to B to C to D./nEach one of these steps requires the intervention of an enzyme, a catalyst that is specialized for that particular step. So, this begins to give you an appreciation of how many distinct biochemical steps one needs in a cell. The numbers probably to make a simple cell, you probably need about a thousand distinct biochemical reactions, each of one of which requires the involvement of an enzyme./nAnd many of these steps, importantly, many of these biochemical steps are endergonic reactions. Where do they get the energy for driving these reactions forward if they're endergonic? ATP. So, the ATP from the energy generating furnace down here is the then spread throughout the cell to power all of these energy consuming reactions. Have a great weekend./nreff: www.mit.edu
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Biochemistry 1 Biology > Introduction to Biology Lecture 2
Video Lectures - Lecture 2/nTopics covered: /nBiochemistry 1/nInstructor: /nProf. Robert A. Weinberg/nTranscript - Lecture 2/nOK. So today we're going to spend a little bit of time on some elementary chemistry just to develop our language that we use with one another. And so when I say hydrogen bond, you don't stare blankly at me and scratch your heads. Many of you have had this already. For many of you this is a review, but it's a useful review. We believe here at MIT of teaching things two or three times often, the same subject matter, but at increasing levels of sophistication./nSo I do this without apology. Our first issue here is how are atoms and molecules held together? And the most familiar way by which atoms and molecules are held together is, of course, the covalent bonds. And covalent bonds have an energy of roughly 80 kilocalories per mole./nAnd that's a rather strong energy to hold together two atoms because the energy, the thermal energy, that is the energy at, let's say, body temperature is about 0.6 kilocalories per mole. And, therefore, if you had a bond, if there was something holding things together that was in this range or two or three or four times higher then the simple thermal energy at room temperature or at body temperature would be sufficient to break apart such a bond./nBut, in fact, this energy, the energy of a covalent bond is so much higher that it's highly unlikely that thermal energy is going to break apart a preexisting covalent bond. And I was just reading yesterday about how people were analyzing the mitochondrial DNA from some Neanderthal bones which were dug up. The last Neanderthal lived around 30,000 years ago, our recently demised cousins./nAnd they were analyzing the DNA sequences. And they got out of those analyses stretches of DNA that were 200, 300 nucleotides long. And that really is stunning testimonial to the fact that under very difficult conditions, nonetheless, complex biological molecules are able to survive over astounding periods of time, indeed those that are held together by the covalent bonds like this./nOf course, you remember the film Jurassic Park where they used PCR reaction to resurrect the DNA of dinosaurs. That's a bit of a fantasy since dinosaurs left us, I guess, about 150 million years ago, something like that. There's a big difference, obviously, between 300,000 and 150 million year ago. Now, the fact is if you look at the way that molecules are actually hooked up, for instance, let's look at a water molecule here./nIdeally there should be no charge on this molecule. And, in fact, there is no net charge. But the truth of the matter is, if one wants to get frank, that oxygen molecules, and we always are here, that oxygen molecules have a greater affinity for electrons than do hydrogen atoms, i.e., they are electronegative. And, therefore, what this means is that the swarms of electrons that are holding all this together at the orbitals are drawn more closely to the oxygen and the hydrogen atoms, i.e., the protons are relatively willing to give up their electrons./nAnd what this means is that there's an unequal distribution. And, as a consequence, there is a fraction of a negative charge here at this end of the molecule and there are fractions of positive charges here because it's not as if they've totally given up the electrons, but the electrons are shifted more in this direction./nAnd this molecule is therefore called a polar molecule by virtue of the fact that here it has a positive pole and here it has a negative pole. There are other pairs of molecules which are relatively equally electronegative. For example, here, if we have a carbon and a hydrogen, these two atoms are roughly equally matched in terms of their ability to pull electrons away, one from the other. And, as a consequence, there is no net shifting of charge. And keep in mind that this delta I show here is only a fraction of an electronic charge. It's not the entire electronic charge moved over./nBut this has important consequences for the entire biochemistry that we're about to get into both today and on Monday. Important because polar molecules, such as water like this, are able to dissolve certain compounds. And nonpolar molecules, which have large arrays of these kinds of bonds or carbon-carbon bonds, these are relatively insoluble in water, and that has important consequences for the organization of biological membranes./nWe might have a carbonyl bond here, that is a C going to an O via a double bond. And here we have, once again, a situation where the oxygen is far more avid in terms of its willingness and interest in pulling electrons toward itself. And, therefore, the carbon gives up a little bit of the electron cloud and it becomes slightly electropositive. Whereas, the oxygen atom becomes slightly electronegative./nNow, the fact of the matter is that there are also other bonds that are noncovalent and are much less energetic. For example, let's talk for a moment about a hydrogen bond. And it's perhaps easiest to demonstrate a hydrogen bond by looking at the structure of two neighboring water molecules in a solution of water of all things./nAnd, the fact of the matter is, let's say we draw one water molecule down here and one water molecule down here. What will happen is that this oxygen atom over here by virtue of its electronegativity will have a certain affinity for pulling this hydrogen atom toward itself. And, in fact, what actually happens in real life, whatever that is at the molecular level, is that this hydrogen atom may actually be bouncing back and forth between these two oxygens./nIt may be rapidly an interchange between them. This interchange causes a strong association between two neighboring water molecules. And, indeed, represents the reason why water does not vaporize at room temperature because the water molecules have a strong affinity or an avidity for one another. And, therefore, just to take some illustrations out of the book, this is the way it's illustrated in the book./nProbably good to have a screen down. And here you can see the way that water molecules are actually arrayed in water. This is the lower illustration here. Just to indicate to you that the hydrogen atoms are not really the possession, the ownership of one molecule of water. They're just constantly being exchanged back and forth. And this back and forth exchange, this sharing of a hydrogen atom is what enables a hydrogen bond of roughly 5 kilocalories of energy per mole to hold things together./n5 kilocalories is not much. It's only one order of magnitude above 0.6 rather than being two orders of magnitude. And, therefore, if one raises the temperature to the level of boiling, if the temperature is high enough, the thermal energy is high enough to rip apart these kinds of associations./nNow, if we were to go back here to look at this carbonyl atom we would find the following sort of situation. Here we have this unequal sharing of electropositive and electronegative bonds. Let's put an acidic group like this. This is a carboxylic acid right here. Here we see a carbon bond to a hydroxyl here via this oxygen atom./nHere, once again, we have an electronegative atom. And, in fact, if we talk about an ionized acid, normally in the absence of ionization there would be a net zero charge right here. But at neutral pH it may well be the case that the association, for various reasons, between this oxygen and this hydrogen will allow the hydrogen, or rather the proton, the nucleus of the hydrogen atom to just wander away. And, therefore, we can imagine there could be a net negative charge here./nA whole, this has one full electron, electronegative charge here, the charge of one electron, and this proton will have ionized, will have left the carboxylic group in which it originated, and now we have an ionized acid group. Either before or even after this ionization, there is a strong affinity of the carboxyl group with the water around it because let's look at what happened before the ionization occurred./nThis carbon here is strong and electronegative. And, therefore, it will participate in hydrogen bonding to the water solvent here, i.e., this proton will be shared a bit between the oxygen of the water molecule and the oxygen right here. Similarly, here this oxygen will be slightly electronegative for the reasons I've just described. And here, once again, there may be some weak hydrogen bonding going on./nAlthough, not as effective as over here where we have a double-bond where we have a lot of concentration of a cloud of electrons pulled towards the oxygen atom. And this begins to give us clues as to why certain molecules are soluble in water and others are insoluble. For example, if we look at aliphatic compounds. Let's look at a compound that's structured like this./nI guess most people would call this pentane. And we can call it that, too. And this has no electronegativity or positivity by virtue of the equal affinities of these two kinds of atoms, that is the hydrogen and the carbons for electrons. And as a consequence, this will not be able to form any hydrogen bonds with a solvent around it if the solvent happens to be water./nSo there's not good bonding here. And this will, in fact, also if one puts this in a solution of water, this will cause all the water molecules to line up in a certain way, almost a quasi-crystal around the aliphatic molecule. They'll be ordered in a certain layer around the aliphatic molecule without being able to form any strong hydrogen bonds with them./nAnd this ordering represents a loss of chaos, a loss of entropy. Entropy is chaos. It's disorder. It's what happens, let's say, at 10:55 when we all leave the room, all of a sudden order becomes chaotic. And here, before this lining up occurred, the water molecules were chaotically arrayed throughout the solvent. After this lining up occurred there was a loss of entropy, there was a loss of chaos./nAnd thermodynamics tells us that generally the ordering of molecules is disfavored. And consequently we now have two reasons why this molecule doesn't like to be in the midst of water. First of all, it's unable to form hydrogen bonds with the solvent. And second of all there is a decrease in the entropy, in the chaos that occurs when this molecule directly confronts water. And because of those two reasons it turns out that this molecule doesn't like to be in water./nThe aliphatic molecule, as one would call this in organic chemistry, doesn't like to be in water. And a dislike of water is often called its hydrophobicity, or we often call it hydro, might as well spell it right, hydrophobic, i.e., it really hates to be in water. In fact, class, there's a second meaning for hydrophobia, or hydrophobic has a second meaning./nEvery five years I ask a class to see who knows what the second meaning of hydrophobia is. This is really obscure. Sorry? Rabies, right. The TAs aren't allowed to answer that. If somebody has rabies, at one stage of rabies, almost near the terminal stage, the individual becomes hydrophobic because he or she doesn't like to drink water, for reasons that are obscure at least to me./nNow, conversely, molecules that have carboxyl group on it would be called hydrophilic. And, as we'll see over this lecture and the next one, these hydrophobic and hydrophilic tendencies tend to have great affects on the overall behavior of molecules. Let's, for example, imagine a situation where we have a long aliphatic tail like this. In fact, these tails can go on in certain aliphatic compounds./nThey can go on for 20 or even 30 carbons. And at the end of this, let's just put arbitrarily a carboxyl group. And let's say we ionized it. So here's an acidic group that's ionized. It's shed its proton. It's actually acquired a negative charge. And now we have something, this molecule is a bit schizoid. Because on one end of it, it loves to be in water, the other end of it hates to be in water./nAnd this has strong affects. It's sometimes called amphipathic, but we don't need to worry about that word. And, therefore, this carboxyl head loves to stick its head, to immerse its head in water. And these things, the aliphatic portion hates to be in water. Now, as a consequence of these rather conflicted feelings that these molecules have about water, we can ask the question what happens when we put such molecules actually into water?/nAnd what we see here is the following. That if we were to construct, for example, a molecule of the sort that has here, in this case we're talking about a molecule that has two hydrophobic tails. We'll get into its detailed structure shortly, but just imagine for a moment two long hydrophobic tails out here ended with a hydrophilic head./nAnd under such situations, if we put thousands of these or millions of these molecules into a solution of water, what we will then see is, no pointer? All right. Pointer? All right. What we will then see is that the hydrophilic head groups, which are here depicted in red, will point their way outwards, they will want to stick their heads in water./nAnd conversely the hydrophobic tails fleeing from the water will actually associate one with the other. And so you have a structure that's called, in this case, an a micelle where you form this little globular sphere where the lipid tails are tucked inside. And, therefore, are actually being shielded from any direct exposure to water. This structure down here, the lipid bilayer, is actually, as we will discuss in greater detail shortly, the overall topology of the way most biological membranes are organized./nIn fact, virtually all of them. Why is that? Because biological membranes separate two hydrophilic or two aqueous spaces. Thank you, sir. A gentleman you are. So here is an aqueous space and here is an aqueous space. And as we see the hydrophilic heads are immersed or sticking their heads into the hydrophilic space./nThis is called a lipid bilayer. And, obviously, it's highly effective for separately these two aqueous compartments. In eukaryotic cells, as I mentioned last time, there is an enormous premium placed on separating and segregating different aqueous compartments which is invariably achieved through the device of constructing these lipid bilayers. Here's a vesicle. A vesicle is more complicated than a micelle./nBecause if you look at the membrane lining the vesicle, you see it's actually a lipid bilayer, but one that in 3-dimensional space is actually a sphere. And in the case of this vesicle, we can well imagine that on the inside of the vesicle water is kept, can be stored, and on the outside of the vesicle water can be stored. And many of the membranes that we see within the cytoplasms themselves are actually constructed on this kind of design./nSo when we draw, for example, in this case the Golgi apparatus, which I mentioned to you in passing last time we met, each one of these membranes here, it's obviously drawn as a double line, but whenever you see a membrane indicated, implicit in that drawing is the fact that each one of these membranes is actually a bilayer. There are never any monolayers of lipids in living cells. Each one of these vesicles you see here is actually a lipid bilayer with an aqueous inside and, once again, aqueous on the outside./nAgain, much of the thermodynamic stability that allows these vesicles to remain intact rather than just diffuse apart is created by these hydrophilic and hydrophobic forces which tie such molecules together or will rip them apart. Now, in truth there are yet other kinds of forces that govern the affinity of molecules to one another. For example, let's imagine a situation where we have an ionized acid group of the sort we just talked about before./nNow, by the way, here, let's say I'll draw the negative charge on one of these two oxygens, if you can see that. But the truth is that the electrons are swarming back and forth, and so the negative charge is shared equally, the negative one electron charge is shared equally between these two oxygen atoms. And this is obviously an area of great electronegativity./nIndependent of that, let's imagine up here we have a basic group, let's say an amine group over here. And, the fact of the matter is, amine groups, NH2 groups, that's what an amine is, here's an amine group. This is a carboxylic group. And the amine group, which is used very often in biochemistry, actually has an affinity. It has an unpaired set of electrons on the nitrogen, and so it likes to attract protons to it, which makes it, causes it to be called basic./nAnd this attraction, the scavenging of protons, perhaps from the water, will obviously give this whole group here a net positive charge, a charge equal to the charge of one proton. Here, once again, we can imagine this is hydrophilic because this charge group can once again also associate quite intimately with aqueous solvent./nNow, independent of any other forces that might exist here, indeed one could imagine situations where there is a sharing of a proton. And, therefore, a hydrogen bond formed between these two. Independent of that is the simple electrostatic interaction of these two groups. That is the mutual attraction of positive and negative groups, one to the other. And the electrostatic interactions, you cannot quantify exactly how many kilocalories a mole there is because the energetic value in electrostatic interaction is equal to one over r squared where r is the distance between these two charged groups./nAnd obviously the further apart you get the weaker the attraction with one another. There are also what are called van der Walls interactions. There are largely of interest to a very small community of biochemists. You probably will never, you may never hear this term again in your life./nAnd van der Waals interactions come from the fact that if we were to have, for example, two molecules over here which are not normally charged in any way, let's just talk about two aliphatic chains again. And I won't put in all the protons and everything, but just imagine a situation like this. What will happen is that because of the fluctuations of electrons, because the electrons are swimming around here all the time, moving from one area to the next they're never equally distributed homogenously over a long period of time, there will be brief instance in time, microseconds or even nanoseconds when there happens to be more electrons over here than right here./nJust by chance. And this area of unequal distribution of electrons will in turn induce the opposite kind of electron shift in a neighboring molecule down here. Obviously, depending on the distance between them./nBut the negative here will repel electrons down here. The positive here will attract electrons down here. And so you will have these two quasi-polar arrangements here and here, very ephemeral, that is lasting for a very short transient period of time. But, nonetheless, sufficient to give a very weak interaction between these two molecules which may persist only for a microsecond and then be dissipated because the charges then redistributed once again./nAnd, as a consequence of that, one has very weak interactions which, in the great scheme of things, play only a very minor role in the overall energy which holds molecules together. Now, with that background in mind, let's begin to elaborate on it, on how we can make molecules that have interesting properties that enable them, among other things, to participate in the construction of lipid bilayers, which will be the first object of our attentions today in terms of actual biochemistry./nSo here's a fatty acid. We see that up here. I, in effect, drew you the structure of a fatty acid up here already once before. And what we can see is through a linkage known as esterification we can create this molecule. So what do I mean by esterification?/nWell, in this case we're talking about a situation here where we have a carbon atom over here like this with a hydroxyl group. You see it over here. And what we're doing is we're dehydrating this, we're pulling out one net molecule of water. And each time we do that, on three separate occasions, what we end up doing is to create instead of this is to create a covalent bond between these two./nAnd so the end product of dehydrating this, pulling out one net molecule of water is that we end up with a structure that looks like this./nAnd you see that happening on at least three different occasions, here, here and here. Well, actually, I should put a carbon over here./nSo here we have three esterifications. The hydroxyl group in each case is reacting with a carboxyl group here pulling out one water, and each case creating what's called triacylglyercol or triglyceride. Triglyceride refers to the fact that we started here with a glycerol and we have now esterified it. Now, in fact, there are two directions here in this kind of reaction./nEsterification is the kind of linkage that we just showed here. And the truth is that vast numbers of biochemical linkages are made by esterification reactions and reversed by reactions that are called simply hydrolysis. And, in this case, what we're referring to is the fact that if one were to reintroduce a water molecule into each of these three linkages, one, two and three, we would break the bond and cause this entire structure to revert to the two precursors that existed or preexisted prior to these three esterification reactions./nAnd time and again you'll see, over the next weeks, that esterification reactions are important for constructing different kinds of molecules. Now, the fact of the matter is we can do other kinds of modifications of a glycerol like this./nHere what we've done, instead of adding a third fatty acid, note what was done here. Here through an esterification, let's look up at this one here, instead of adding a third fatty acid, we've saved, we've reserved one of the three groups of the glycerol. Here's what we saw just before. We've saved one of the three groups of the glycerol and put on instead this highly hydrophilic phosphate group, once again through a dehydration reaction, an esterification reaction./nAnd now what we've done is add insult to injury because in the absence of this phosphate it would have a hydroxyl here which is mildly hydrophilic. But now look how strongly charged this is. Here are two negative charges, one electron each. And this is already a bit electronegative. So here we have an extremely potent hydrophilic entity. And here the degree of schizophrenia between one end of the molecule and the other is greatly exaggerated. Here, in fact, this is extremely hydrophilic./nAnd, as a consequence of that, this really likes to stick its head inside water. And when we therefore talk about, we draw the images of different kinds of membranes, like this I showed you before the two tails. Here you saw the two tails I drew before in that diagram. Here's what we can imagine they actually look like in more real molecular terms. And the hydrophilic heads sticking in the water, this is just repeating what we saw before, become even more hydrophilic if we look at a molecule like this./nLet's look at this thing here. Here's a very long hydrophobic tail. Here are the two glycerols once again. Here is the phosphate. And keep in mind that phosphate obviously has these extra oxygens. Phosphate can react with more than just one partner, the glycerol down here. In this case we've added this group up here. And this group up here is, once again, this happens to be a serine which is an amino acid, this also happens to be quite hydrophilic./nHere's our old friend the basic amino group. Here's the carboxyl group. This is a bit hydrophobic, CH2. And then we once again have the hydrophilic head here. And, therefore, we imagine, if we look at what's called a space-filling model, and a space-filling model really is intended to show us what one imagines if one had this vision, which we don't have, how much space each of these atoms would actually take up if one were able to see them./nAnd here we see this space filling model. This lipid molecule here is actually slightly kinked with its hydrophilic head tucked into the water space. And so here's actually the way that many biological membranes look in terms of the way that they are constructed. Now, the fact of the matter is this also affords the cell the ability to segregate contents on one or the other side of whatever lipid bilayer it happens to have constructed./nAnd here we can see about the semi-permeability, how permeable these membranes are to different kinds of molecules. Permeability obviously refers to the ability of this membrane to obstruct or to allow the migration of molecules from one side to the other./nIons, and these ions we see right here are obviously highly hydrophilic by virtue of their charge. That's explains, in fact, why, for example, table salt goes so readily into solution, because it readily ionizes into sodium, NA and CL, which then are avidly taken up by the water molecules. So these are highly hydrophilic ions. And the questions is, can they go from one side of the membrane to the other? And the answer is absolutely not or highly improbably. Why?/nBecause these are so highly hydrophilic, the water molecules love to gather around them and form hydrogen bonds and electrostatic bonds with them. And if one of these ions ventures over here, it's going from an area where it's warmly embraced by the solvent molecules to an area where these molecules intensely dislike these ions. And, therefore, thermodynamically the entrance of any one of these ions into the membrane, into the hydrophobic portion of the membrane is highly disfavored, which makes the membrane essentially, for all practical purposes, impermeable./nThe same can be said of glucose which happens to be a carbohydrate. We'll talk about it shortly. But it's also nicely hydrophilic. It also can go in water. In fact, it can go through. And it's actually the case, to my knowledge, that one doesn't really understand to this day why lipid bilayers are reasonably permeable to water. You would say, well, water shouldn't be able to go through./nIt clearly doesn't have to have a net positive or negative charge, but the physical chemist, if you asked them why does water, why is water able to go through lipid bilayers? They'll say, well, we've been working on that and we'll get you an answer in the next five or ten years. And they said that 40 years ago and 30 years ago, and they're still saying it. And we don't really understand why water goes through, which is an embarrassment because here's one of the fundamental biochemical properties of living matter that is poorly understood. Gases can go right through./nAnd amino acids, ATP, glucose 6 phosphate, highly hydrophilic, can also not go through. Now, the advantage of this is that a cell can accumulate large concentrations of these molecules either on the inside or it can pump them to the outside. In other words, it can create great gradients in the concentrations of different kinds of ions. For example, in many cells, the concentration of calcium, CA++ is a thousand times higher on the outside of the cell than on the inside of the cell which is a testimonial to how impermeable these lipid bilayer membranes are./nThe fact of the matter is I'm fudging a little bit here because in the lipid bilayers of the plasma membrane of the cell, the outer membrane of the cell that we talked about in passing last time, there are ion pumps which are constantly working away pumping ions from one side to the other overcomes the little bit of leakage which may have occurred if a calcium ion happens to have snuck through in one direction or the other./nAnd we end up expending a lot of energy to keep these ion gradients in appropriate concentrations on the outside and the inside. In fact, virtually all the energy that is expended in our brain, almost all of it is expended to power the ion pumps which are constantly insuring that the concentrations of certain ions on the outside and the inside of neurons are kept at their proper respective levels./nIt could therefore be that actually more than half of our metabolic burden every day is expended just keeping the ions segregated on the outside and inside of cells. For example, potassium is at high levels inside cells, sodium is at high levels outside cells, just to site some arbitrary examples. There are also, by the way, as I mentioned last time, channels./nAnd channels are actually just little doughnut shaped objects which are placed, inserted into lipid bilayers in the plasma membranes and just allow for the passive diffusion of an ion through them, through the doughnut hole enabling an ion, so if here's the lipid bilayer, not showing its two things, these kinds of doughnut shaped protein aggregates will allow the passage of ions in one direction or another./nAnd here energy is not being expended to enable this passage. It may just be through diffusion. If there's a higher concentration of ion on side of the lipid bilayer and a lower one on this side, this diffusion will allow the ion to migrate through the bore of the ion channel from one side to the other. In fact, even though this does not involve the expenditure of energy on the part of the cell, the cell may actually use a gating mechanism to open or close these channels./nWhen the channels are closed then the ions cannot move through. When the channels are gated open then diffusion can take over and insure the transfer, the transportation of ions from one side to the other. Now, having said that, we can begin to look at yet other higher level structures. Here, by the way, is a better drawing than the one I provided you. This comes from your book of what a vesicle looks like./nHere's what it looks like under the electron microscope and here's what it looks like when a talented rather than hapless and hopeless artist like myself tries to draw it. So let's just say that's our intro into lipids and membranes. And let's move onto the next layer of complexity. And the next layer of complexity in terms of molecules represents carbohydrates./nAnd when we talk about a carbohydrate amongst ourselves we're talking about a molecule which, roughly speaking, has one carbon atom for every water molecule. And we'll shortly indulge ourselves in talking about all kinds of different carbohydrate molecules. Here is really one of the most important carbohydrate molecules, glucose. And what should we note about glucose? Well, the first thing you should see is that glucose has six carbon atoms. And, therefore, as a consequence it's called a hexose./nWe're going to talk about pentoses very shortly. They only have five, to state the obvious. Glycerol, which we talked about before, is also considered in one sense a carbohydrate, but it's been called by some people a triose. It only has three carbon atoms. And you can imagine, therefore, in principal that there are certain biochemical mechanisms which indeed exist which enable one to join two glycerol molecules, one to the other, to create something like a hexose, glucose./nIn fact, what we see from this drawing, expertly drawn by yours truly, is that the hexose molecule isn't really a linear molecule in solution. What happens is that because of various steric and thermodynamic forces it likes to cyclize. So let me just mention, I've just used two words that are useful to know about./nSteric or stereochemistry refers to the 3-dimensional structure of a molecule. And, obviously, the stereochemistry of a molecule is dictated by the flexibility with which participating atoms can form bonds, whether we have a trivalent atom like nitrogen or a tetravalent atom like carbon or a monovalent like hydrogen./nAnd these structures, the stereochemistry is dictated both by what atoms are present here and by thermodynamic considerations which cause this particular hexose, indeed virtually all hexoses, to cyclize. When I say cyclize, obviously I mean to form a circular structure. Here we note one thing. You can see how the hydroxyl here actually attacks the positively charged carbon here in order to form this cyclic structure./nYou see one of the six points on this hexagonal structure here is oxygen. It's not carbon at all. So there is one oxygen and five carbons. And one of the carbons is relegated, is exiled to outside of the circle. It's sometimes called an extracyclic because it's sticking out from the actual circle./nAnd this is the structure in which glucose actually exists inside cells. And, in fact, there is, in truth, two alternative ways by which glucose can cyclize, whether the oxygen attacks the carbon on the carbonyl group underneath or on top. And you see that gives us two alternative structures. What's different about them? Well, if we think about this hexose as existing in a plane, or the hexagon is in a plane/nIn this case the oxygen is above the plane and the hydrogen is below the plane. With equal probability you can have these two atoms reversed where hydrogen is now above the plane and hydroxyl is below the plane. And both of these structures, these alternative structures can fairly be considered to be glucose. Now, let's get a little bit more complicated. Here we have fructose and we have galactose./nAnd what we see here is, by the way, that we have exactly the same number of carbon atoms and hydrogen atoms and oxygen atoms but they're hooked up slightly differently. And here now we begin to get very picky about the disposition, the orientation of these different kinds of hydroxyls and hydrogens. And note, by the way, here that in many cases one doesn't even put in the H for the hydrogen. It's just implied by the end of this line./nAnd here, if you were to look at this, you'll see here now we have two extra cyclic carbons. Here's galactose which is yet another hexose. These are all hexoses, but their stereochemistry creates quite different kinds of structures. And it turns out that this stereochemistry is extremely important. These molecules function very differently, one from the other./nAnd, for example, to the extent that glucose is used in different kinds of energy metabolism and to the extent that galactose is not, there must be certain biochemical mechanisms in which one has catalysts, the catalysts that we call enzymes that ensure that one can convert one of these hexoses through an enzyme into, let's say a less useful one into a more useful one, glucose, which can readily be burnt up by the energy-generating machinery. Here we've gone yet another order of magnitude more complex because we've gone from a monosaccharide, i.e., one or another hexose, to a disaccharide./nAnd here's common table sugar. And here you see that it's formed once again through an esterification reaction, i.e., there is a dehydration reaction between this hydroxyl here and this hydroxyl here. And biochemists take the orientation of these hydroxyl and hydrogen groups very seriously. Now, you can say they're a bit obsessive. Indeed they probably are./nBut, nonetheless, we can admit that the specific orientations of all these things dictate very importantly the difference between here, in this case sucrose, and in this case lactose. Why is this important? Well, this is the sugar in milk sugar. This is the dominant sugar in milk sugar, lactose. And half the world, as adults, cannot absorb this. All kinds of unpleasant things happen when they actually drink milk./nHow many people here are lactose intolerant? It's nothing to be ashamed of. I'm married to a very lactose intolerant person. She's otherwise very nice. The fact is that the enzyme to break down lactose, it's an enzyme which is called lactase. And here we have yet another nomenclature item. So lactase is the enzyme which breaks down lactose./nAnd, by the way, this is just the harbinger of many other enzymes we're going to talk about in the future that end in A-S-E. Whereas, carbohydrates, many of them end in O-S-E, as you've already sensed. So it turns out that the enzyme lactase is made in large amounts by most mammals very early in life. Why? To be able to breakdown the milk sugar that comes in their mother's milk./nBut once mammals are weaned there's no reason on earth for them to continue to make lactase, in their stomach for example. And, as a consequence, in most mammals the production of lactase is shut down later in life. And for some weird quirk of human history, a significant proportion of humanity has learned how to retain the ability to make lactose through adulthood. And, as a consequence, people can go and have ice cream until the age of 70, 80 or 90 without becoming very bloated./nAnd we don't need to get into all the details, but you can begin to imagine. And what happens is, therefore, the lactase enzyme is shut down in their stomach. It depends. Sometimes they lose it at the age of 10 or 15 or 20. And then, for the rest of their lives, whenever they have a milk containing product, in fact, my son is also lactose intolerant. I'm surrounded by these people. Again, he is otherwise a tolerant person but he's lactose intolerant./nSo this lactose molecule will go into the stomach, it will remain undigested, it will remain a disaccharide instead of being cleaved into two monosaccharides. The two monosaccharides are no problem because they can readily be interconverted. The galactose can be readily converted into glucose, and glucose is the universal currency of carbohydrate energy. And so this disaccharide passes through the stomach unaltered and it gets into the intestines, in the small intestine and the large intestine./nAnd it turns out we have more bacterial cells in our gut than we have our own cells in the rest of the body. Imagine that. And there are a lot of bacteria that are waiting around in the gut for just a little gulp of lactose. And they never get it because most people break down their lactose long before it gets into the intestine. But here we have these lactose intolerant people./nThe disaccharide gets into the gut and the bacteria go to town. They've been waiting around for years, decades for a little bit of lactose. And now it finally arrives and they go to town, ad they start metabolizing it and they ferment and they produce lots of gas and other kinds of byproducts. And, as a consequence, this makes people very uncomfortable. Just to show you, now, the fact is that lactose intolerance people can perfectly well break down sucrose, obviously. This is one of the great energy sources from plants./nBut they cannot break this down. And I emphasize that point to indicate that the stereochemical differences between different kinds of carbohydrates makes a very important difference. An enzyme like sucrase will break down the sucrose but it will not touch lactose. So there's a high degree of stereospecificity as it's called in the trade. Here we now go to another step forward that we're going to pursue in much greater detail next time./nBecause here, for the first time, we talk about polymerization. We're making polymers. Where the large number of hydroxyl groups on these monosaccharides affords one many opportunities to make very long linear aggregates end-to-end like this or even side branches. If you imagine that each one of these hydroxyls, in principle, represents a site for possible esterification, i.e., the formation of a bond to a neighboring side chain./nHere we see these two linear chains and here we see the branch which is afforded, which is made possible by the availability of these unutilized hydroxyl side chains which are just waiting around to participate, if the opportunity allows them, in some kind of esterification reaction to form a covalent bond. Here is, by the way, glycogen, which is the way we store a lot of sugar in our liver./nHere's a starch, which is what we get from many plants. And here's another very interesting polysaccharide. It's called cellulose. And we cannot digest cellulose, but termites can. And why they can is something we'll have to wait until next time to learn about. Have a great weekend. See you on Monday.
Tags // Biochemistry 1 mit
Added: March 31, 2009, 6:51 pm
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