 Good morning. I'm glad to see some of you decided to come back. I'll start having office hours today. So at 9 o'clock, I'll go from here down to 2084 VLSB. It's not a very big room. So if you want to get a seat, you should try to get there early. And office hours will be 9 to 10 today. And then tomorrow morning, I will have office hours from 9 to 10. Not many people come on Tuesdays and Thursdays. So that's just a little tip. OK, I'm going to hopefully finish talking about protein structure today and then be able hopefully to get on to lipids. So on Friday, I talked a bit about proteins. I told you what a protein is. A protein is a polymer made up of amino acids. And then we talked about the 20 are called natural amino acids. These are amino acids that are found in proteins. And at the end of the hour, and I wanted you to try to focus on the different structures of those R groups, their hydrophobicity or their hydrophilicity. And you will see today how this comes into play in relation to the structure of proteins, the final three-dimensional structure of protein. And at the end of the hour, I showed you how you can take two amino acids, condense them to form a dipeptide. And I put another reaction like that on the board. In this case, I have one amino acid. It is aspartate. A second amino acid is alanine. And one gets from this aspartyl phenylalanine. Now the point of this is, again, to show you how you are condensing these amino acids. And then at the carboxyl end, you would continue to add amino acids up to the thousands. This is a packet of equal. Equal is an artificial sugar. It's a non-sucrose-containing sweetener. Equal contains a compound which is known as aspartame. Aspartame is a modified form of aspartyl phenylalanine. It's not exactly the same. You make this, and then you do some modifications. And you get a compound that is, I don't know, 200 times sweeter than sugar and doesn't have the problems of sugar, high sugar levels in one's blood. And the last time I looked, aspartame was a $100 million product. The moral of this is there's money to be made in biology. But I think you probably know that. So maybe that's why some of you are going into biology. So today, we're going to start talking about the different levels, the way protein structure is discussed. There's a hierarchy of structure within proteins. And in this case, I will be referring to the figures, figure 10, 11, 12. There's no 12, 13, and on. The first way people talk about proteins, the first level that is discussed is what is called the primary level, primary structure of a protein. The primary structure of a protein is the linear sequence of amino acids within that protein. So you put in aspartate, you put in phenylalanine, you continue to put in amino acids. Where does this sequence come from? It comes from the genetic code. It's coded for in your DNA. We will not talk about that now because Bob Fisher will talk extensively about that. This is a schematic of the structure of insulin. The structure of insulin is actually given in figure 13. Insulin was one of the first proteins where the primary structure was deduced. And that was done by a biochemist Fred Sanger at Cambridge University in England in the 1950s. And insulin is a small protein. It has about 60 amino acids. And there are a couple of points I want to make about it. You can see the complete amino acid sequence of the protein in figure 13. There's a polypeptide A and a polypeptide B. These are often referred to as subunits when you have two different polypeptides in a protein. They are called subunits rather than polypeptides. The protein has a disulfide bond. I talked about last time the amino acid cysteine can form disulfide bonds. There's an internal disulfide bond. And there's a disulfide bond between the two chains. So it's a rather simple protein, but it actually has a rather complicated structure. Sanger not only determined the complete amino acid sequence of these two proteins, but he worked out all of the techniques for doing amino acid sequencing. This was in the 1950s when basically you had to do chemistry and the chemistry involved taking off one amino acid at a time and identifying it and then putting it back together. Sanger received the Nobel Prize for that work and then subsequently he received the second Nobel Prize, one of very few people to ever receive two Nobel Prizes. So the linear sequence of amino acids in the protein is its primary structure. Incidentally, no one determines the sequence of an amino of a protein this way anymore. Get into that. OK, the next level of structure, the secondary structure. Oh, here's my protein. I brought my protein in today. So here's my protein. And what you're doing in the primary structure is you're putting amino acids together to get a linear chain. The second level of structure within protein is that there are regions within the protein where folding occurs. So regional folding. So within this thing, it doesn't stay as a long linear chain. It may form, for example, if I could get it, an alpha helix. So I'm making a little coil in this region. The folding within the secondary structure of a protein is stabilized by peptide bond hydrogen bonds. This is what's shown in figure 11. You can see there's a polypeptide chain running in one direction, a polypeptide chain running in another direction. And there are hydrogen bonds between peptide components that stabilize the secondary structure of a protein. There are two major structural elements that are considered to be part of secondary structure. One is an alpha helix, which is what I tried to show you here, shown in figure 11 the bottom. And the second is something called beta pleated sheet, where you have a strand of amino acids going in one direction, interacting with a strand of amino acids going in another direction. These two components were proposed by Linus Pauling. Has everybody heard of Linus Pauling? Yeah, you didn't know he did this. Linus Pauling was probably one of the greatest chemists of the 20th century. He worked on the nature of the chemical bond. And his major contribution in chemistry was in protein structure, proposing that alpha helical regions and beta sheet regions exist within proteins. Of course, he went on to greater or more notorious fame by proposing everybody should be eating five grams of vitamin C a day. OK, remember, the key word in terms of secondary structure is regional or localized. So within a protein, there's a small region where you can form either a beta sheet or an alpha helix. The third level of protein structure, tertiary structure, is the final folded protein structure that exists. So I may have a protein that has a bit of alpha helix in it like this. This thing folds up to a final three-dimensional structure. That's not very pretty, but I think you can get the idea. The stabilization in this structure is through our group interactions, shown in figure 10. The order here is crazy, but I'm giving you the number. Our group interactions mean side-chain interactions, the side chains of the amino acids. For example, this carboxyl group in the side chain of aspartate can interact with an amino group side chain. That would be considered as a, well, it's not exactly, an ionic bond. Dysulfide bonds are side-chain interactions. Hydrophobic interactions between the nonpolar amino acids are side-chain interactions. Many of these bonds are weak. Hydrogen bonds are weak. Ionic interactions are weak. These disulfide bonds are not weak. These are covalent bonds, very strong bonds. So they basically lock a protein into a structure. The structure that a single polypeptide or a single subunit protein has is called its native conformation. I've put that on the board up here. The final folded structure is denoted its native conformation. And this is the structure in which the molecule is active. So many proteins, which only have one single polypeptide chain, fold into their tertiary structure. That's the act of form of that protein, activity. What do I mean by activity? Well, we have enzymes, which react with substrates. We have antibodies, which react with antigens. We have receptors, which react with molecules that may be going in or out of a cell. All of those are biological functions, which are driven by or through the native conformation of a protein. For what we call soluble proteins, soluble proteins are proteins that are found in the liquid phase of the cell. We'll talk about this probably by the end of the week. Most of these proteins take on a conformation, which is called globular. Basically, they form a ball. And I'll talk in more detail about some of the forces that are involved in a protein folding. How does a protein fold? That remains a very substantial question in biochemistry, modern biology. There are also proteins, many proteins, for example, that contain more than a single polypeptide chain, more than what we would say a single subunit. Here's an example here. Insulin has two different polypeptides, chains in it, an H chain and a B chain. These proteins have what is called tertiary structure. So any protein that has tertiary structure has more than a single polypeptide chain. However, the forces that stabilize tertiary structure are identical to those that stabilize, I'm sorry, quaternary structure are the same as those that stabilize tertiary structure. There are group interactions. So you may have, like you have in insulin, you may have one chain cross-linked, essentially, covalently bound to another by disulfide bonds. You may have hydrophobic regions that are involved in stabilization of these interactions. Figures 14 summarize what I've put on the board and what I've just said. It shows you, again, primary, secondary, tertiary, quaternary structure, the stabilizing forces that occur within them, and there's a little schematic in B that shows how these proteins are folding. You have to understand this. You have to understand the different levels of protein structure, what they mean and what's involved in stabilization of the various forms. Now, the question that biochemists have been working on for a very, very long time, since the 1960s, essentially, is how do proteins fold? What are the forces that allow a protein to go from this to this? And it still isn't fully understood. What is known is that the information for protein folding is contained in the primary structure of a protein. What do I mean by that? I mean, if you look at a protein's primary structure, that dictates somehow the final three-dimensional, the native conformation of that protein. And that idea came about by a very clever experiment done by Anfenson, Chris Anfenson, in the 1960s, working with an enzyme called ribonuclease. You don't have to know anything about ribonuclease at this point other than it's an enzyme. Substrate is RNA, ribonucleic acid. And the experiment he did was ribonuclease has eight SH groups, eight of these cysteine residues, eight SH groups. And these eight SH groups form four SS bonds. Four disulfide bonds. Chemically, you can reduce these SS bonds to SH bonds. And that he did. That's basically what is shown in figure 15. You have disulfide bonds. They're reduced to sulfhydyls. When you do that, this enzyme loses its activity. Why does it lose its activity? Because you have altered its native conformation. If you alter the native conformation of an enzyme, even a little bit, it probably will lose its activity. So the native conformation is biologically active, but altered conformation will usually not be active. So Anfenson produced an inactive protein by reducing the disulfide bonds that are present in this protein. And then he took away whatever reagent he used to go from this to this. And he let the protein refold. If there are four disulfide bonds from eight SH groups, I don't remember what the number is. Maybe it's 64, maybe it's 16. There are a number of ways these four disulfide bonds can form, just randomly. This is a random process. Three and six can react, or three and 14 can react, et cetera. And presumably, those forms of the enzyme would be inactive because they don't have the same SS bonds that are in the native enzyme. And when he examined his product from the enzyme that was refolded, it had 100% of the activity as the native enzyme. So somehow, once the amino acid sequence is determined, that sequence that has within it the information for folding and producing the biologically active native conformation of the protein. This experiment was really the one that led to that conclusion. Now, I have to tell you one additional thing because many of the things we say in here, they're not 100% true, should I say that, or something like that. Or they may have been true last year and they're not true this year. That's the way biology is. The idea that all the information for proper protein folding is in the sequence of the amino acid is not fully true. It is now realized that there are proteins, which are known as chaperones, whose function it is to assist in the proper folding of proteins. And how chaperones actually work is not fully understood. But it's clear, to me at least, that protein folding is so important that the cell has even devoted specific proteins to assist in the proper folding of these proteins. Chaperones particularly are involved where there's a quaternary structure of proteins where you have to bring different subunits together to get them into the final active conformation. So you should just be aware that it's not quite as simple as Anfinsen let us to believe. But still, that's the way it is sometimes. The other interesting feature of Figure 15 is it also illustrates the point that one can take a protein and one can denature it. There is a process which is called denaturation. And that's what Anfinsen did. If you take a protein and you disrupt its active conformation, it has become denatured. And you might expect denatured proteins are inactive. And from what I've just said, under some conditions, they can be reactivated. There are various physical methods that are used to denature proteins. The most common one is take proteins and heat them. You heat a protein up to boiling. It aggregates, it forms big clumps that look like chewing gum and it precipitates out of solution. That is irreversible, but it certainly is also inactivating the protein. There are various chemicals. One can add organic solvents. Generally will inactivate proteins. Sometimes they can be reactivated. But if one denatures a protein, I'd say the rule is generally that that protein is losing its function. So we come to this big point that structure is related to function. That is, if you have a structurally active protein and you alter that structure, you are probably going to alter the function of that protein. If you look at figure 17, and here's a case where I can't draw this. Figure 17 represents the structures of some proteins. Now, what I think one of the wonders of modern biochemistry is over the past 50 years, is the development of techniques that have allowed us to determine the complete three-dimensional structure of proteins at the molecular level. Actually, the initial impetus for doing this work came in the 1930s when a biochemist named Sumner was the first person to actually crystallize a protein molecule. People did not think you could crystallize proteins because they're too big. And Sumner crystallized urease, an enzyme that breaks down urea. And 50 years later, people took, were able to take crystals of proteins and determine the complete three-dimensional structure of the molecule. Can I have the overhead up? Great, thank you very much. Okay, what one does in determining the structure of a protein is one does it, uses a technique which is called x-ray crystallography. You take your protein crystal and you shoot x-rays at it. Now, if it's a crystal, the assumption is it's very ordered because things don't crystallize unless they're ordered. And you shoot x-rays at your crystal and the x-ray bounces off, depending on the organization of the atoms within that molecule. And you get what is called a diffraction pattern. This is a diffraction pattern, x-ray diffraction pattern for a molecule known as myoglobin. And in the 50s, myoglobin is a rather simple molecule. It's brother, you probably are more aware of, hemoglobin. Both of these proteins had been crystallized. And in the 1960s, two British biochemists, John Kendrew and Max Perutz worked out the three, the complete structure of these molecules. And from this, now how you go from this to this, which is the structure of myoglobin, is something I can't explain to you, okay? But you guys are probably smarter than I am. What's shown in this figure is the polypeptide chain. You don't have the complete protein. The complete protein looks like this. These are all of the atoms in the protein. So what is shown in this figure is the polypeptide chain and you can see that there are regions in this protein which are helical. There's one, two, three, there may be more behind, at least four alpha helical regions. So myoglobin is a protein which has a lot of secondary structure and it has a lot of alpha helices. Does not appear to have many beta sheets in it, okay? If you wanna see it in living color, there's living color, much prettier actually. When this was done, myoglobin and hemoglobin were the first proteins for which three of these structures were obtained. Now, you can go online to various data sets and you can find hundreds of proteins which have been crystallized and their structure is determined. It is no longer an esoteric thing to do but in the 1960s when the first structures came out for these molecules, people were sort of blown away. Myoglobin is a protein that has a molecular weight of about 15,000 and it has only a tertiary structure. Hemoglobin, much more complicated structure. If you look in figure 18B, because hemoglobin is a molecular weight of about 64,000, it has a quaternary structures. It has two alpha chains and two beta chains. So it contains two different proteins, subunits. These alphas and betas have nothing to do with the alpha helix and the beta sheet. We use the same name sometimes, it's confusing. But it's a much more complicated structure because it's a much more complicated protein but these two proteins were the first proteins for which we have complete three-dimensional structures. This has given us an enormous amount of information. If you know the structure of a protein at the level of detail that you get from these kinds of analysis, you can start to understand the interactions that are holding the protein together which is one of the questions that still remains unanswered. How do proteins fold? You can start to look at active sites of enzymes, where things bind within proteins. Both hemoglobin and myoglobin are oxygen-carrying proteins, right? You know about hemoglobin. And they are both red proteins. They are red because in addition to having amino acids in them, they both contain something which is known as heme and we'll talk about heme later. Heme is what I call a prosthetic group. Its structure is given in figure 14C. It's a rather complicated structure. It's a molecule that has about a lot of nitrogens in it surrounding an iron atom. That's why hemoglobin and myoglobin are red. That's why hemoglobin and myoglobin carry oxygen because the oxygen binds to this heme group. And we will talk, I'm just introducing this to you now, we will talk in much more detail about prosthetic groups when we talk about enzyme structure and function. Jumping around in the figures, so bear with me. If you look at figure 17, I said these were three-dimensional structures, x-ray structures of various proteins. And the only reason I wanted to give you these pictures is to show you that, for example, structure A, which is a viral protein from tobacco, mosaic virus, that's a protein that is predominantly alpha helical, as is myoglobin. Myoglobin, as I said, if you look at figure wherever it went, 14C is the structure of myoglobin, there are many alpha helices in that protein, but no beta sheet. If you look at the next protein, which is immunoglobin, that is a structure, those arrows, those long arrows represent beta sheets. So if you see an arrow going this direction, that means the amino acids are going like this, and then coming in this direction, it means there are hydrogen bonds between these two regions of amino acids. This protein is predominantly beta sheet, and if you look at the next one, which is an enzyme, hexokinase, we'll talk about that enzyme later, it's kind of a mixture. It's got a little alpha helix, it's got a little beta sheet. So there's no real rule that you can follow to say this is going to be this, and this is going to be this. But as people determined more and more structures of proteins, it became clear that there are something within proteins which are called domains, there are regions of proteins. If you have a protein, for example, that I'll make it very simple. Let's say it is a protein that somehow interacts with a magnesium ion. This is probably too simple, but it's okay. Magnesium plus plus. And you compare the structure of a lot of proteins that contain, that interact with magnesium, they will generally have the same structure in that region. So what you see in figure 18a is a protein that as I think you can see from the picture of that, there's a top region on this protein and there's a bottom region. And the top region is a region that interacts with a substrate and the bottom region is a region that interacts with a component required for the reaction of that substrate. And again, if you start to compare the structure of various proteins that interact with these kinds of molecules, they may start to show you the same regions, the same domain. So many proteins that would, for example, interact with magnesium would have a similar domain. Nature is conservative, that's what this tells me. Once you've built it and it works, there's no sense rebuilding it. So there is some order to all of this. Okay, the best example I can give you about structure is related to function also is a story about Linus Pauling. There is a disease called sickle cell anemia where people have very, very low red blood cell count and when they looked at the blood of these people under a microscope, instead of having these nice donut shapes, red blood cells, the red blood cells were sickle shaped. And the name came, was given sickle cell anemia. These sickle cell anemia patients don't carry oxygen as well as normal people and they have a shortened life because of this. And I think it was in the 40s or 50s. Pauling came along and he was interested in proteins by then and he said sickle cell anemia is a molecular disease. I don't know if anybody knew what that meant. In 1945 to 50, I'm not sure they're making, what's he talking about? Well, molecular disease. When people could determine the amino acid sequence of proteins, they did the sequence of the hemoglobin and normal people and sickle cell hemoglobin and it was discovered that in one of the chains, I don't know if it's the alpha or beta, to be perfectly honest, one amino acid residue, residue six, was a glutamic acid in normal hemoglobin and was valine in sickle cell anemia patients. Those who had sickle cell hemoglobin had the substitution of a valine for a glutamic acid. If you go back and look at your amino acid table, you will see that glutamic acid is a very, very polar amino acid. It's got a carboxyl group on it. It's basically like a smartic acid. Like this except there's another CH2. Valine is a very hydrophobic amino acid, one of the most hydrophobic amino acids. So this one change in amino acid sequence was sufficient to alter the function of the hemoglobin in these cells, change the shape of the red blood cell because there was some clumping of these hydrophobic regions in the sickle cell anemia hemoglobin and it misshaped the red blood cell. And this is given as the sort of classical example of how structure is related to function. If you change the structure of a protein, i.e. you substitute a valine for a glutamate, you are gonna change the activity, the function of that protein. Now later on, we'll talk more about this subject and when Bob Fisher talks about mutations, you'll hear again about it. This is essentially what a mutation is. A mutation is the alteration in the sequence of DNA which produces an altered protein and most mutations are bad. Structure is related to function. Okay, are there any questions? Because I'm done with proteins. Everybody's with me. Okay, look at the figures. They're, as I said, the figures are much better than the things I can draw on the board. I've got enough time to talk about lipids. Lipids are the next group of biological molecules that I wanna refer to. Lipids, again, are not polymers. They don't have the same property that polysaccharides and nucleic acids and proteins have. They're a group of molecules that have one thing in common and that is, what can I erase? They are insoluble in water. So if somebody mentions a lipid, that's the first thing you should remember. So they are very hydrophobic and there are three sort of groups of molecules that we'll talk about. One are fats. The second are phospholipids and the third are steroids, some steroids plus some miscellaneous compounds. Okay, what are fats? If you look at, we're actually missing a page. Page 10 is missing. Does everybody have page 10 or is it just me? It's after page 11. Oh yeah, it's after page 11. They're in the wrong order. Okay, let's do page 10, much more important. Okay, fats contain two components. They contain glycerol, which is a three-carbon alcohol, very simple molecule. I can even drug glycerol. That's glycerol. So there are three OH groups in glycerol and it contains what are known as fatty acids. Fatty acids are shown in the left-hand part of figure 19. Fatty acids are long, basically hydrocarbons. If you look at the compounds in figure 19, for example, palmitic acid, which has 16 carbons. There's a carboxyl group at one end. That's why it's an acid. Everything else is basically hydrocarbon. And this is why fats are not soluble in water because they have hydrocarbons running through them. There are two kinds of fatty acids. There are saturated fatty acids such as palmitic acid. Every carbon has two hydrogens on it. There are unsaturated fatty acids such as stearic acid where there should be, I'm sorry, oleic acid, where there's a double bond in the middle of the molecule. The structure of those two fatty acids is different. The saturated one, like stearic acid, is basically a straight chain. The double bond produces a kink in the middle. So if you look to the left and you look at oleic acid, you have in one case just something that looks like this and in the other case you have something that looks like this. That's where the double bond is. To get a fat, you take glycerol and you esterify it with fatty acids. That is what is shown in figure 20. An ester and a carboxylic acid is an ester. So you get either one, two, or three fatty acids attached to a glycerol molecule. If you have one, you have a monoglyceride. If you have two, you have a diglyceride. If you have three, you have, you got it, okay? Triglycerides. So those are fats, very, very sort of simple molecules, but important. The fatty acids that are found in fats can vary. They can be 14, 16, 18 carbons. It really depends on the organism. It depends on plants, animals there. There's a fairly large range of fatty acids that can be found, but this does not affect the general properties of these molecules. It is now recognized that animal fats which contain fully saturated fatty acids, like lard, okay, are not good for you, right? Everybody knows that now. Because they are related to heart disease. Whereas the kind of fats that are found in plants which have a high amount of unsaturation seem to be better. Fats in plants are really oils. Olive oil, everybody loves olive oil now. Every house has big bottles of olive oil, uses it for everything. Canola oil, we use canola oil. Like olive oil, but cheaper. They're oils because at room temperature, these molecules are liquids. Whereas at room temperature, animal fats are solids. And this is one of the problems with the animal fats. This relates to the structure of the fatty acids, whether they are straight chains or whether they have these kinked structures because the kinks produce less packing. The molecules are, the side chains are further apart and they are, this yields the product of them being oils at room temperatures. So I don't have to give the lecture about why you should have unsaturated fats versus saturated fats because everybody understands that. So these are the fats, okay? The second group that I mentioned are phospholipids. Phospholipids are more important and we will be talking more about phospholipids. Phospholipids shown in figure 21 have three different components. Firstly, they still have a glycerol, they still have fatty acids, but they will only have two fatty acids and the third OH group in glycerol is attached to a phosphate group. It's a sterified to a phosphate. Phosphate is basically phosphoric acid. So it's an acid, it reacts with an alcohol and you get a phospho ester bond. That's what's shown in figure 21 and so that those are the first several components. The phosphate group is also attached to something other than the glycerol. It's attached to a low molecular weight organic alcohol, such as shown in figure 22. So you have, if you look at figure 22, the first compound at the top of that is the amino acid serine. Well, the amino acid serine has an OH group in it, so it can a sterified to the phosphate group which is a sterified to the glycerol. So we have a phosphate group which is this and one of these OHs is a sterified to glycerol and another of these OHs is a sterified to a organic alcohol, such as serine, but also there's a variety of compounds. Ethanolamine, choline, glycerol, anacetol, on and on and on and on. So a lipid, a phospholipid looks kinda like a fat but it has a dual identity. It has a region, the way this is generally drawn is it has a head group which is polar and it has two fatty acid side chains. The polarity of the so-called head group comes from the presence of the phosphate group because a lot of OHs in there and also the organic alcohol which is also a hydrophilic compound. So this tends to be a polar region, this tends to be a non-polar region. So phospholipids have this sort of identity that's two fold, polarity, polar region, non-polar region. If you take phospholipids and you throw them into solution they spontaneously form what is known as a phospholipid bilayer. So this is in solution, phospholipid bilayer is formed and that is shown in figure 23. So what you can see in figure 23, each one of these represents a phospholipid molecule. They look like this and I'm not gonna say much more about this because we will talk in detail about this. Why do these things form spontaneously? This is a region which is very hydrophobic, strong interaction between fatty acid side chains. Do you expect water to be in there? No water, there's no water in this region. Water's out here and over here because there's interaction with the polar head groups of the phospholipid molecule. So you have a funny kind of molecule. There's a polar region which is in the water, the aqueous environment. There's a non-polar region from which water is excluded. I'm not gonna talk more about these because when we talk about the structure of biological membranes, you will see in some detail that the phospholipid bilayer is a key component in biological membranes. Okay, I've got enough to finish this. The last group of molecules that I talk about, again, these are not soluble in water, fats are not soluble in water. Steroids, if you look at figure 24, there are a number of steroid molecules like cholesterol, testosterone, all of those kinds of molecules. And you look at the structures of those molecules, they're basically pure hydrocarbon. They're not soluble in water so we characterize them as lipids. There are also carotenoids which are found in certainly in plants which, whose structures are shown in the middle of that figure, again, highly non-polar molecules, very hydrophobic and for that reason, they're under miscellaneous compounds because they're not soluble in water. And another one, which most people wouldn't consider a lipid, but if I'm gonna call it a molecule that's not soluble in water, chlorophyll, which we don't have, but plants have various forms of chlorophyll. Again, they have a very, very hydrophobic tail of carbon atoms, which makes them virtually insoluble in water and they would be a miscellaneous compound which is also characterized. If you've gotta give it a name, call it a lipid because it's not soluble in water. Let's see, there's anything else I wanted to say. No, oh, let me just say a word about trans fats. Well, we're on fats and I have a minute or two. The other furor in the world now is trans fats. Trans fats are also compounds which have recently been discovered to be not very good for you. Trans fats are produced when chemists want to take, to make liquids solid because liquids at room temperature are much harder to store than solids. This is why margarine was developed. So what they did was they started to add hydrogens across double bonds and they discovered that the hydrogens can go in in two directions. They can be on the same side, which is cis, or they can be on different sides of that double bond, trans. And the evidence now seems to indicate that trans fats are really not very healthy and there's been a big push just recently, the last year or two, to get rid of trans fats from all foods or get them down to very, very, very small levels. So there is some importance in lipids in our health and happiness, okay? Okay, on Wednesday I'll talk briefly about carbohydrates and nucleic acids and then we're gonna talk about cells.