BIO 2B03 Module 1, Lecture 1 PDF

BIO 2B03 Module scripts Module 1, lecture 1 Script Notes Slide 1 Hi everyone, and welcome to BIOLOGY 2B03- Cell Biology. As a blended learning course, BIO2B03 Is course that will cover a broad array of interconnected...

BIO 2B03 Module scripts Module 1, lecture 1 Script Notes Slide 1 Hi everyone, and welcome to BIOLOGY 2B03- Cell Biology. As a blended learning course, BIO2B03 Is course that will cover a broad array of interconnected themes. This will include considering how proteins are made, their structure, their function , and for those who do so… how some proteins exit the cell. We will also explore a number of ways that a cell is able to perceive signals from each other and their environment, and how the response to these signals often results in short term or long term cellular changes. Finally, we will explore how cell division is a very controlled and regulated process which eventually results in mitosis and the division of one cell into two identical daughter cells. This is a process that’s especially important for many tissues in our body. But, all cells also have a shelf-life. During times of stress, damage or when triggered by pre-programmed cellular signals, a cell will undergo self-destruction through a process called apoptosis But before we get started with these interesting topics, we have to start with basic principles. Today, we will get started with our first Lecture for Module 1, From polypeptide to protein, where we will explore our different amino acids and their chemical interactions. Let’s get started! Slide 2 The objectives for this module are to review the principles of the primary structure of a polypeptide, to take an overview of the of the properties of amino acid variable side chains (or R‐groups), and to describe the chemical interactions essential to protein structure. Slide 3 What is a protein? What are the functions of proteins? I want you to take a moment just to pause and think about your definition of a protein and think about what proteins do in the cell. You might’ve come up with the definition of proteins such as a string of amino acids, a polypeptide chain or apolymer of other small molecules. When you think about the function of proteins, we can basically answer that by saying proteins do everything. Some examples you might have come up with could include the fact that Proteins are structural components of the cell, (tubules, actins and cytoskeletal elements) that proteins are sensors for environmental change and that they may help with mechanisms for relaying information from the outside of the cell to the inside of the cell, initiating changes in cell behavior. You may also remember that proteins are enzymes that can metabolize chemical reactions. Or that proteins can regulate gene expression (acting as activators or silencers that allow or inhibit gene expression) Proteins could also be modifiers that change the function of other proteins, turning proteins on and off. We also know that proteins are signals that are produced by cells, and are required for signaling between cells. And of course, proteins are also receptors that receive those signals, and they are the components of the transduction pathways that interpret those signals. Or maybe you were thinking, proteins form molecular motors, in that they are able to transport components around the cell. Or that proteins are important for the unique identity and function of membrane‐bound organelles in the cell. All of these functions are true, and all functions outline the importance of proteins in all of our cells! Slide 4 How can one type of molecule accomplish all of this? Unlike RNA and DNA, which assume a limited number of structures, there is a virtually an unlimited number of molecular structures that proteins can assume. Each protein has its own unique, defined structure that enables it to carry out a particular function. If we look at the structure of proteins we could see that there is a diverse array of what proteins can look like: diversity in size and diversity in structure. The shape and structure of a protein is ultimately going to specify function of the protein. We see here, some examples of enzymes such as glutamine synthetase that forms a large donut‐shape structure with an active domain in the middle. We see smaller globular proteins such as the signal insulin or the oxygen‐ carrying protein complex hemoglobin. We also see here, the typical Y shape of the immunoglobulin protein that is necessary for recognizing antigens, viruses and bacteria, in the cell and removing them. Finally, we see a smaller enzyme over at the right hand side, adenylate kinase, containing a structure that is essential for defining the substrate binding domain and the activity domain for this enzyme. Slide 5 Proteins are composed of amino acids. The linear array of amino acids is the primary structure. We see here, 5 amino acids that have been strung together to form a small polypeptide. One amino acid residue in the chain is circled in green. Each core central alpha carbon is shown with the associated variable side chain. In this primary sequence, these variable side chains are R1 R2 R3, etc from left to right or from N‐terminus to C‐terminus- that is, amino end to carboxyl end. Connecting each of these amino acids together to form a chain ofamino acid residues are peptide bonds. The peptide bonds are represented here by the orange line between a carboxyl group and amino group. We have a total of 20 possible different amino acids that can be incorporated into newly synthesized polypeptides or proteins. We will see later that even after a polypeptide is formed, post‐ translational modifications can alter the properties of individual amino acids, and as a result, the properties of a protein, as a whole. Slide 6 A single amino acid is represented here. The 20 possible amino acids all have the same basic structure. The central alpha carbon has four side chains: a hydrogen side chain an amino group a carboxyl group and thevariable R group. It is the variable R group that is going to define the property of each amino acid residue, but it is the accumulated R groups that are going to define the properties of a polypeptide or protein. Slide 7 Since the the R-groups or side chains of amino acids are essential to understanding the properties of proteins, we need to think about how these side chains differ from one another. We will look at the properties of the individual side chains. But keep in mind that what is important are the emergent properties that come from the interactions of all the side chains in a protein. These interactions will determine how the protein folds and it will determine the characteristics of the whole protein. The amino acid side chains differ in their size, shape, charge, hydrophobicity, and reactivity. Cell biologists classify these amino acid residue side chains into groups based upon their solubility in water or the polarity of the side chain. Slide 8 Solubility is a physical property of a molecule that refers to its ability to transiently interact with water through hydrogen bonding. This is thermodynamically favorable. So- if hydrogen bonds can form with water, the molecule can be soluble. If a molecule cannot form these hydrogen bonds, then it is insoluble. A soluble protein typically carries hydrophilic amino acid residues on the external surface. In contrast hydrophobic molecules are not polarized and cannot form these hydrogen bonds, so …water repels these molecules in favor of bonding with itself. Slide 9 What sorts of molecules are hydrophobic? Really, anything that is unable to form a hydrogen bond. Typically we think of oil or fats that do not mix with water. Now, why do they not interact with water? Well, because these oils or fat contain saturated hydrophobic long carbon chains called hydrocarbons. We can then extend this to our amino acid residues. We can look for those amino acid residue side chains that have these types of hydrocarbon chains. Hydrophobic amino acids have types of non‐polar side chains and are water insoluble, or perhaps only slightly soluble. Because of this, these amino acids tend to be found in the interior of cytosolic proteins and form a hydrophobic core. But keep in mind, that a protein found in a hydrophobic environment (such as a protein that’s embedded in a membrane), for these proteins, they will have the opposite structure with the hydrophobic amino acids accumulating on the exterior of the protein, facing towards the phospholipid tails of the membrane layer. Slide 10 We can divide the hydrophobic amino acid residues into 2 categories: aromatic amino acids and aliphatic amino acids. Phenylalanine, tyrosine and tryptophan have hydrophobic aromatic rings in their R-groups. These aromatic rings are very hydrophobic. Note that as cell biologists, we have placed tyrosine into this hydrophobic category, yet it does have a hydroxyl group on the end of its R-group, a group that is capable of forming a hydrogen bond. So really tyrosine falls into both categories, both hydrophobic and hydrophilic. At the right, we have our 5 hydrophobic, aliphatic amino acids. Notice that the R-groups for these amino acids have long hydrocarbon chains. Alanine has just one methyl side chain, valine isoleucine, leucine, and methionine all have longer hydrocarbon chains. Notice also that methionine has a sulfur within its chain. Overall, amino acids with aromatic or aliphatic R- groups (or side chains) will be classified as hydrophobic. Slide 11 What about hydrophilic amino acid residues? Hydrophilic molecules are water soluble, and for that reason we find our hydrophilic amino acid residues on the exterior of soluble cytosolic proteins. Water soluble molecules have portions that are charge- polarized and capable of forming hydrogen bonds. Remember this is relevant to our cells at a physiological pH- a pH of 7. So we’re looking at molecules with a hydroxyl group or a charged oxygen at pH 7, or molecules that have this amino group with a positive charge at a pH of 7. Slide 12 Here we see examples of amino acids that carry charged side chains. On the left, amino acids with positively charged side chains, lysine and arginine. On the right, amino acids with negatively charged side chains, aspartic acid and glutamic acid. These four amino acids are hydrophilic, and are prime contributors to the overall charge of a protein or a domain of a protein. Slide 13 There are also uncharged, polar hydrophilic amino acid residues. Shown here are the amino acids threonine and serine, with polar hydroxyl groups that can participate in hydrogen bonds, and asparagine and glutamine that are uncharged but have these polar amine groups. Slide 14 Finally, we have a collection of special amino acid residues. Cysteine is a special amino acid, because it is able to form covalent bonds, through its sulfur atom, with other cysteine amino acids. These covalent bonds are called disulfide bridges or cysteine bridges Glycine is special because it is very small. The side chain is a single hydrogen. The small size of glycine allows for it to tuck in small places in a folding protein and allow for bends in a polypeptide chain. Proline is special because it’s R- group forms a covalent bond with the amino group of the amino acid. This can lead to a kink or bend forming in a polypeptide chain, that is essential for the structure of many proteins And finally, we have our amino acid  histidine, which has an amino diethyl side chain, that shifts between a positive charge and neutral charge, depending on the pH of the environment For our course, make sure you review all of the properties of the amino acid side chains. We will not ask you to draw the structures of these amino acids, but you should be able to recognize and know the unique properties of all 20 amino acids. Knowing these properties will be essential for us to understand the unique structure and function of different proteins. Slide 15 To make a protein, amino acids must be covalently bound together through peptide bonds. A peptide bond is formed by a condensation reaction, which simply means that water is released during the formation of this covalent bond. Here we have two amino acids side by side. We can see the carboxyl group of the amino acid on the left and the amino group of the amino acid on the right. Through a condensation reaction, one molecule of water is released and a peptide bond is formed. Slide 16 Protein chains are typically depicted from left to right, from N‐terminus (or amino end) to C‐ terminus (or carboxyl end). Protein synthesis takes place during the process of translation. During this process of translation, new amino acid residues are added to the carboxyl end. As a result, the N‐ terminus of any protein is the first amino acid in the chain and subsequent amino acids are added to the carboxyl end, that is, the right hand end of the growing chain. Slide 17 Translation happens in the ribosome. Shown here is a video animation of the process of translation from the Howard Hughes Medical Institute’s collection of BioInteractives. Slide 18 Video, no script Slide 19 Let’s think a little bit about how we get from a polypeptide to something that is actually functional in the cell. Seen here, on the left hand side we have a representation of a polypeptide string, a linear array of amino acids that is not functional yet in the cell. We need a three‐dimensional protein such as the one on the right that has a series of loops and bends and sheets that define structural and functional domains within the protein. The structure on the right is dependent upon the amino acid sequence that we see on the left. But-- it is not simply that amino acid sequence that is going to allow us to get to our three- dimensional structure. We are going to see that the polypeptide has to be modified in various ways in order to allow for this folding to occur. Slide 20 There are four levels of organization to get from a sequence of amino acids to a functional 3‐ dimensional protein. The primary structure is that linear array of amino acids. Slide 21 The primary structure of the polypeptide is the linear array or sequence of amino acids and is determined by the sequence of nucleotides in the coding DNA. This image represents the processes of transcription and translation that lead to the synthesis of a polypeptide chain. To start, a sequence from the double‐ stranded DNA helix is transcribed into a single‐ stranded messenger RNA. That messenger RNA is processed to remove introns and produce a mature messenger RNA, which essentially contains only the codons that are required for the synthesis of the protein. The mature messenger RNA molecule is then exported out of the nucleus into the cytosol of the cell, where it is translated by the ribosome. So, the amino acid sequence is determined by that nucleotide sequence in the original associated gene. The number of different polypeptide sequences is limited by two factors: 1) there are 20 distinct amino acids that can become incorporated into a polypeptide, 2) the number of amino acid found in that polypeptide. So for any polypeptide string that is“n” amino acids long, the number of different arrangements of amino acids in a chain would be defined by 20 to the power of “n”. Given this, we can see that there are infinite number of polypeptide sequences that could be produced in a cell. Slide 22 Polypeptides fold spontaneously and assume what is called a random‐coil structure. A random‐coil is best described as a periodically ordered structure of the protein. Proteins might not have a single stable structure, but a collection of related structures that they switch between. The term statistical coil is a representation of this idea and suggests that the protein spends most of its time in a particular structure, but not 100% of its time. This structure may be stabilized by interactions within the polypeptide, but also interactions with other proteins and other molecules as well. The structure that the polypeptide assumes most of the time, or native structure, is the functional protein structure. The local interactions that maintain protein shape are for the most part non‐covalent interactions that include ionic bonds and hydrogen bonds, Van der Waals forces and what is called the hydrophobic effect. Individually, these are weak attractive forces, but in their aggregate they form a very strong association, and provide stability to the folded protein structure. Slide 23 Let’s step through each of these interactions. First of all, the ionic bond. This is an attraction between a positively charged cation and a negatively charged anion. Represented here in this image, is the interaction between glutamate and lysine, where we have a positively charged cation on the lysine side chain anda negatively charged anion on the glutamate side chain. If we look at this in the context of a simple polypeptide, we can see that this ionic interaction is assisting in holding the two arms of the polypeptide together in this shape. Now one bond isn’t going to be enough to do that, but we’re going to see how an accumulation of these interactions can maintain protein shape. Slide 24 A hydrogen bond is the interaction between a partially charged hydrogen atom in a molecular dipole, such as what we see in water, and an unpaired electron from another atom. Represented at the bottom right hand corner here is a polarized covalent bond, where there is not a full charge, but apartial charge or dipole moment. In a water molecule, oxygen has a partial negative charge, a negative dipole, whereas the hydrogen is exhibiting apartial positive charge, or a positive dipole. This can happen not only in water molecules, but in other covalent bonds as well. Represented here as well, is a hydrogen bond that is forming between the molecular dipoles at different side chains of two distinct amino acid residues. The hydrogen bond contributes here to protein shape Slide 25 Hydrophobic effects are the aggregation of nonpolar molecules in aqueous medium, simply to reduce the number of interactions with water. An example of this would be an oil droplet in water. All of the hydrophobic molecules aggregate together to minimize the surface area that is exposed to the hydrophilic aqueous environment. The aqueous environment of the cytosol can similarly induce these hydrophobic effects for hydrophobic amino acids in proteins. Slide 26 A Van der Waal’s interaction, which is also known as London Dispersion forces, is a weak non‐specific attractive force. It results from a transient dipole that is induced when two atoms are very close together and they perturb the distribution of electrons in each other. This transient dipole only exists when these atoms are close to one another. The association that occurs though is not a strong one, it is not as strong as a hydrogen bond or ionic bond, but ifwe accumulate many of these Van der Waal’s forces together, these interactions can be quite strong together. Slide 27 Let’s take a look at an organismal example of Van der Waal’s interactions – the amazing ability of a gecko to walk on walls and ceilings. How is this possible? At the bottom of this slide, we see a close‐up of the gecko’s foot. If we take an even closer look at the surface of its feet (as we see on the right), we can see structures called setae.These setae are small little fibers that are present on the surface of these feet. Each one of these little setae induces a transient dipole with the underlying surface causing a Van der Waal’s interaction. Now one seta would  not be enough to hold a gecko in place, Slide 28 … but when there are millions of these together, all on the entire surface of the foot, we find a very strong interaction. It is the sum strength of all of these Van der Waals interactions that allow geckos to walk along the ceiling. This is a neat observation, because we can use this to model adhesive surfaces. We can do this by looking at nanotubes that are mimicking the setae that we find in gecko feet. Similarly, in proteins, individual Van der Waal’s interactions are very weak attractive forces. But the accumulated effect of many bonds like these, makes for a very strong overall association between two different proteins or within a protein. Slide 29 Folding is the result of interactions between amino acid residuest hat hold a polypeptide into shape. Here is a short animation that shows a polypeptide folding. Before we get started, keep in mind a few things: 1) The color of the spheres and the type of interactions that can occur. Here, red spheres are representing amino acids with hydrophobic chains, purple and blue spheres are representing amino acids that can form hydrogen bonds, and yellow and green represent amino acids that can form ionic bonds. These interactions are important inallowing the peptide to form a three‐dimensional structure. 2) Remember that folding is a random process; bonds are forming and bonds are also breaking as different bonds form. Folding is dynamic. 3) Folding depends upon the number and the strength of bonds and interactions that are maintaining protein conformation. 4) Also, note that for simplicity, in this animation, the arms of the polypeptide move through one another, that would be like us walking through a wall. It doesn’t happen in an actual protein, but it is just shown to help us visualize the animation. Slide 30 I n this animation, we start with a short polypeptide. The grey spheres are amino acids, their specific identities do not affect folding. The polypeptide begins to fold and takes on a shape. Again, keep in mind that this is a random process. The polypeptide might unfold, but then it might fold again. The hydrophobic side chains in this polypeptide are going to aggregate together in a hydrophobic core, that hydrophobic core is also important in stabilizing the  protein. Remember that hiding those hydrophobic side chains is thermodynamically favorable. We see bonds beginning to form, blue and purple form a hydrogen bond that is holding those arms together in that particular conformation. Yellow and green form an ionic bond, holding together the arms. And hydrophobic red spheres are aggregating predominately in the core of the protein. All of these bonds are collectively contributing to a stable structure for the protein. Slide 31 That’s all for Module 1, Lecture 1. In summary, what we hope you’ve taken away, is that our understanding of the chemical properties of the amino acid side chains allows usunderstand the underlying principles of protein folding, and is especially important as we will see in the course, for protein function!