BISC 307L 2nd Edition Lecture 32 Current LectureImmunoglobulinsIf you take serum proteins, they are all globular proteins – so they are called globlulins. And if you separate these globulins by electrophoresis, you come up with five classes. First class, while globular, is not called a globulin. It is the A class – A stands for Albumin, the most abundant serum protein. The other 4 classes are called globulins: alpha1, alpha2, beta, and gamma. The gamma globulinsare antibodies or immunoglobulins. There are 5 classes of immunoglobulins: these are all protein secreted by B-lymphocytes. The most abundant in the plasma is IgG, which is shown in the box. It has four parts, and two pairs of identical subunits – two pairs of light chains and two pairs of heavy chains joined by disulfide bridges. Has a Y shaped structure. There are constant C and variable V regions.The places that bind the antigen are the tips of the variable regions. The heavy chain has a hingeregion, which is flexible. That allows the angle between the two antigen binding sites to vary and move.Fab fragments occur when you take antibodies and you digest them with a proteolytic enzyme called papaine. It cleaves to make two fragments, the antigen binding fragment (Fab) and the crystallizable fragment (Fc). Called the crystallizable fragment because these Fc’s are made up largely of a chunk of the constant region, which is pretty constant between antibodies, so you can put all these Fc’s together and crystallize them. All of these are secretions of plasma cells, or activated B lymphocytes. How do plasma cells get activated? At the top, you can see IgD. And its structure looks like the thing in the box, but thereis a transmembrane region in the constant region of IgD’s. So IgD is a membrane protein. It is anchored by the transmembrane region, and an IgD is the B cell receptor. When an antigen binds to the antigen bindin region of that IgD, this B cell will get activated. When the B cell is activated, it proliferates to form plasma cells and memory cells. The plasma cells will secrete antibodies, and the first ones they secrete will be IgM, which is a huge molecule consisting of 5 subunits IgD held together by disulfide bonds and a joining chain that connects a couple of them. It is too large to leak out of capillaries. Even in inflamed capillaries, which are much more leaky, IgM does not leak out. IgM’s arena action is mainly in the blood. So after about a month of a plasma cell secreting IgM, it begins to undergo class switching. And what happens is that the gene for the constant region is switched form the type that is specific for IgM to the type that is specific for IgG First class switch is from IgM to IgG. The same plasma cell that was secreting IgM now secretes IgG. The class switching was in the constant region, and not the antigen binding region. We are going to see that during its embryonic development, each B cell rearranges its genome and allows mutations to accumulate in the genes for these immunoglobulins – once that’s done, the DNA of that B cell is permanently rearranged for the rest of its life and doesn’t undergo any further changes or mutations. So when a given plasma cell switches from the constant region gene for IgM to one that is specific for IgG, the variable regions don’t change. The part of the DNA that gives rise to the antigen binding regions of protein remains the same. In other words, the IgG has the same identical antigen binding specificity a the IgM did. And for that matter, both of these immunoglobulins have the identical antigen binding specificity as the IgD receptor in the first place. Therein lies the specificity of thesystem – the same antigen that activated the B cell will be the one that is attacked by its antibodies. IgG is a smaller molecule, it can leak out of capillaries somewhat, and leaks out freely in areas ofinflammation. It is the only maternal antibody that can cross the palcenta and get into fetal blood. It helps to confer the mother’s immunity onto the fetus.Some plasma cells, when they switch from making IgM to IgG, they switch to the gene for the IgE constant region instead of the IgG. This is abnormal, and that plasma cell will spew out large amounts of IgE, which will trigger allergies. IgE is associated with allergies and parasitic worm infections. IgA looks like a dimer of the basic subunits. IgA is the most abundant antibody found in mucus secretions. It has a joining chan in pink and a secretory component in blue. How does it get to the mucus? Shown below.Transcytosis of IgAMucus membrane consists of a single layer of epithelial cells. Below the basolateral membrane are the other cells including the plasma cells. IgA antibodies are coming from plasma cells, which are in the tissue below the basolateral membrane of the epithelia. This plasma cell, whenactivated, will secrete IgM and then class switch and secrete IgA, which consists of the two basicsubunits and the joining chain. This binds to a poly-IG receptor on the basolateral membrane of epithelial cells and when this IgA binds to the poly-IG receptor, this triggers endocytosis of the receptor immunoglobulin complex and the whole thing is taken into the cell in a vesicle that gets translocated by transcytosis to the other side and the enzymes inside the vesicle cleave theconnection between the receptor and the membrane. The remainder of that receptor (the orange thing) is permanently bound to the IgA, and that’s how you end up with the IgA dimer and its attached secretory component in the mucus above the epithelium. Rearrangement of Immunoglobulin GenesIf every antibody has a different variable region, how many different types are there relative to the number of genes in the human genome? There are 25,000 genes in the human genome. The antibody repertoire is on the order of a hundred billion, so there is a major mismatch in the numbers. Solution to the puzzle was a mechanism of recombination of the genes for immunoglobulins. On the left side is shown the gene for a light chain in an immature B lymphocyte. The polypeptide light chain itself consists of a variable region and a constant region. The genes for that are located at different places in the undifferentiated DNA up at the top – there is a variable region, a joining region, and a constant region. The exons are separated by the noncolored introns. In addition, since this is a secreted protein, there is a leader sequence