Specificity, Diversity, and Immunoglobulin genes Flashcards
How is the binding of the antibody and antigen?
It is a non-covalent interaction based ionic bonds, hydrogen bonding, van der Waals forces, and hydrophobic interactions between the amino acid side chains of the antibody’s CDRs and corresponding residues on the antigenic determinant.
Explain Cross-reactivity:
This refers to the tendency of one antibody to react with more than one antigen, which has to do with affinity. We say colloquially that an antibody is ‘against’ mumps virus if it was obtained from an donor immunized with mumps, or if it reacts with mumps with a high association constant (high affinity); but we must re member that what the antibody really has is a combining site made up of six CDRs whose position, charge, and hydrophobicity distribution is such that an antigenic determinant of the mumps virus binds it with observable affinity. Other antigenic determinants might also fit it; if they did so detectably, we would say that the antibody cross-reacted with those determinants.
T cell mediated immunity has similar specificity and cross- reactivity. ►The two cross -reacting epitopes may have very similar amino acid sequences. Sometimes they don’t: but then the charge/hydrophobicity distribution will be similar.
Give some examples of cross-reactivity:
-Some antigenic determinants on the lethal smallpox virus are similar to ones on the almost harmless cowpox virus. If you immunize p eople with cowpox, the cross-reactivity with smallpox is such that they will be immune to that, too
Example2: formaldehyde will change incredibly poisonous tetanus toxin into a harmless toxoid , which retains its immuno genicity; if you’re immunized with toxoid, your antibodies cross-react with and neutralize toxin (due to similar antigenic determinant structures).
–>To activate a B cell to produce antibody is a complex process. For purposes of our discussion now, it can be divided into two phases. First, binding of antigen to the B cell’s receptors (membrane-bound versions of the antibody it will eventually secrete) occurs with a particular Ka . If this binding is strong enough, the second step, activation of the B cell, can take place.
►So an antigen which binds with low affinity may never activate the cell; but if another antigen comes along which not only binds but activates, the product of the cell (the secreted antibody) may combine with the low affinity antigen well enough to be inconvenient (due to similar structural composition).
A non-trivial example: The heart valves contain an antigen, laminin, which cross-reacts with Group A streptococci. Obviously, the antigen in the valves does not normally activate the corresponding B cells, or we’d all have an autoimmune disease. When people get a streptococcal infection, the streptococcal antigens do activate these B cells because they bind to them with sufficient affinity. Then the released antibody can react with heart valves; with low affinity, it is true, but occasionally, in some people, with enough affinity to lead to a destructive, complement- mediated, inflammatory process: rheumatic heart disease.
What is the Clonal Selection theory?
Clonal Selection Theory
It said that each cell of the immune system is programmed to make only one antibody (T cells weren’t known yet, but the theory covers them, too); that the choice of which antibody the cell will make is random, not dependent on outside information; and that the entire population of potential antibody-making cells pre-exists in a normal individual, even before any contact with antigens.
When a new antigen is introduced into the body, it comes into contact with a huge number of lymphocytes, and when it encounters one to whose receptors it binds with sufficient affinity, it activates it, resulting in expansion of that clone and production of that antibody.
► The best- fitting clones are selected by antigen. This theory was initially hard to accept, but has now been shown to be true; and it is the central concept in immunology. Clonal selection is Darwinian; survival of the fittingest, as it were.
Allotypic exclusion, how is it important to know what it is?
- The lambda, kappa, and H chain gene families are all on different chromosomes. A potential problem arises because, since we’re diploid, each cell has two copies of each gene, maternal and paternal.Only one H chain (maternal or paternal in origin) and one L chain (either kappa or lambda, either maternal or paternal) are synthesized in any one B cell.
- All the other genes are silenced. Though the person can make two H-chain allotypes, each individual B cell makes only one. It is reminiscent of the way each female cell turns off one or the other of its X chromosomes; and this fact makes our job of understanding how we generate antibody diversity a little easier.
DNA Recombination:
Explain how the Variable domain gene region:
Explain how the Heav chain is made:
How to make a Light chain is very similar to the process of making a heavy chain, what are the differences?
LIGHT CHAINS : L chain gene rearrangement is similar, but they have only V and J segments, no D; and only one C domain gene, kappa or lambda, depending on the cell’s choice .
What are RAG recombinases?
What is the name of the syndrome of missing RAG?
The enzymes that do the recombination of antibody and T cell receptor DNA are called RAG-1 and RAG-2 recombinases.
-The recombinases first bind splice signals to the right of a D segment and the left of a J segment, pull them together, and then cut and splice. Then they look for a splice sequence to t he right of a V segment and do it again.
►If RAGs are knocked out, mice make neither B nor T cells. It happens in humans, too — very rarely (Omenn Syndrome). The RAG gene system appears in evolution with the jawed vertebrates; lampreys don’t have it.
What is the mechanism for generating diversity by somatic variation?
Which enzyme adds nucleotides at random?
What is an N region?
What the price/result of the randomness of the mechanisms?
The V-D and D-J joins are unpredictably “sloppy.” The cell uses randomizing mechanisms:
- First, exonucleases for chewing away a few nucleotides after the DNA is cut but before two gene segments (D to J, V to DJ) are joined.
- Second, for adding a few nucleotides as well, an enzyme called terminal deoxynucleotidyl transferase, TdT, which doesn’t use a template so its additions are random.
What does this mean?
►Thus you can’t predict the sequence at the joining area (► which is called an “N” region ; there is one on each side of the D segment ); it might be obvious that V7 has joined to D2, let’s say, but in this cell there’s an extra alanine and tyrosine there, and in that one there is a leucine missing. This produces a lot more completely random diversity. There is a price for it: two times out of three the N region, being of random length, will create a **frame-shift mutation, that is, a nonsense codon which terminates transcription.
Allotypic exclusion and receptor editing:
ALLOTYPIC EXCLUSION, PART 2 . Because sloppy recombination often ends up with a frame-shift mutation; when one examines a particular B cell, one may find H and L genes that have been abortively rearranged, that is, in such a way as to produce nonsense codons. When this happens the cell tries again with the other allele; if things work, it goes on to become a B cell, if not, complete antibody cannot be made and the cell dies.
-If it gets lucky on the first try, though, it doesn’t try the other allele. So although any one cell is theoretically capable of making 2 H chains (by rearranging both maternal and paternal loci,) and 4 light chains (maternal and paternal, kappa and lambda,) that never happens; it makes only one of each — all other alleles are excluded .
Receptor editing, what is it?. Although we just said that a B cell tries to rearrange each allele just once, that isn’t strictly accurate. In some cells, when a rearrangement is detected as faulty (say a stop codon is generated), or when an anti-self receptor has been displayed, if the recombinases (RAG genes) are still active it can ‘ try again. ’ Sometimes this results in a successful cell. The process is called receptor editing.
How does somatic hypermutation could increase the affinity of the newly made antobody?
What is affinity maturation?
What enzyme allows hypermutation to happen?
It does not happen for T cells, only B cells.
SOMATIC HYPERMUTATION. ► And, finally and surprisingly, another source of receptor diversity :
the recombined V(D)J unit is “hypermutable”; each time a B cell divides in response to antigenic stimulation there is a good chance that one of the daughters will make a slightly different antibody. Selection by antigen of the best-fitting mutants after antigenic stimulation allows a gradual increase of affinity during an immune response — an exceptionally nice design feature called affinity maturation. (For T cells, somatic mutation after contact with antigen does not seem to take place.)
How hypermutation works?
Activation-Induced [cytidine] Deaminase (AID) converts a random cytosine in the CDR gene regions to uracil. So a cytosine: guanine pair becomes a uracil: guanine mismatch. The uracil bases are removed by the repair enzyme uracil-DNA glycosylase. Error-prone DNA polymerases then fill in the gap, creating mostly single-base substitution mutations, so at the end of cell division one daughter may be making a different (worse? better?) antibody.
How does class switching happens?
Do this process requires T-cell help?
If no T cell help, what is the onlt Ig that can be made?
A single mature B cell starts by making both IgM and IgD, which it puts into its membrane as receptors, and then later it may switch to making IgG, IgE, or IgA.
►In all cases, the L chain and the VH domain stays the same but the C region of the H chain changes.
As may be anticipated by now, what happens is that the cell which has put its particular H-chain VDJ combination together with its mu and delta genes (as shown in the diagrams) goes back to its DNA, does a loop-out of mu and delta, and puts VDJ next to the C- region gene of gamma or epsilon or alpha, while excising and discarding intervening DNA.
-This is our second example of DNA recombination. The new mRNA, then, may be VDJα or VDJγ or VDJε. Thus a cell which is making IgM can go on to make IgG, but a cell making IgG cannot go back to making IgM; the mu information is physically gone . ►‘M to G’ or ‘M to A’ or ‘M to E’ switches are pretty much the rule in antibody responses, and require T cell help; without it, only IgM responses are possible.
There are some antigens for which T cell help is not (normally) possible, and the response to them is IgM only.
What enzyme helps with class-switching?
The enzyme AID also is involved in class switching. Thus B cells recombine DNA two different ways with this enzyme.
T cells do neither hypermutation nor class switching
