Lecture 27 - Immunity II Flashcards
What are antibodies?
Antibodies (aka immunoglobulins or Ig) are essential for adult survival and makeup around 20% of the protein in the blood plasma
What is the basic structure of antibodies?
2 antigen binding sites and consists of 2 light and 2 heavy chains
What are the different classes of antibodies?
5 classes - IgG, IgM, IgA, IgD and IgE where each has its OWN HEAVY CHAIN.
Also sub-classes (e.g. IgG1, IgG2, IgG3, IgG4) - each have different hinge and tail structures giving unique characteristics.
What occurs when an antigen binds to the antigen receptor of a B-cell?
Proliferation and differentiation - antibodies are secreted from plasma cells
What are features of IgM?
- IgMs are the first class of antibody that a developing B cell makes and major class secreted during the into blood on first exposure to an antigen.
- IgM is a pentamer composed of five 4-chain units, giving it a total of 10 antigen-binding sites.
- When antigens with multiple antigenic determinants bind to IgM, it alters the structures of the pentamer, allowing it to activate the complement system. This system is a molecular process that is activated by the constant domain that does the killing of the cells.
What are features of IgGs?
- the most abundant antibody found in the body
- made of 2 copies of 2 proteins (4 in total), linked by covalent di-sulphide bonds: 2x Heavy chains (around 440aa) and 2x Light chains (around 220aa)
- each chain consists of variable and constant domains
What does the constant domains interact with?
Other parts of the immune system - e.g. innate immune & complement systems)
What do variable domains make up?
the antigen binding sites
What are both light & heavy chains made up of?
made up of repeating 110aa domains as immunoglobulin domains each of which contains an internal di-sulphide bond. Likely to be the result of gene duplication during evolution
Describe the size of the variable and constant regions of heavy and light chains.
Variable region - same size for light and heavy & chains
Constant region larger in heavy chain than light
What does the antigen binding region consist of?
2 variable domains made up of a single modified Ig domain and containing 3 hyper variable regions.
What is the Ig domains that make up the constant part of the heavy chain encoded by?
A single exon
How many antibody molecules can a naive, unchallenged human immune system generate?
1 x 10^12 (a primary antibody repertoire)
What can a mature immune system do?
Can make antibodies able to bind to essentially any antigen essentially infinite flexibility - this is able to occur through generation of antibody diversity
How many genes does the entire genome encode?
25,000
How does generation of antibody diversity occur?
- The Ig domains that make up the constant part of the heavy chain are each encoded by a single exon.
- the DNA encoding the variable domain is a bit more complicated.
- in the germ line a k-light chain gene variable domain contains 40x V domains, 5x J domains and a single C domain
- and a heavy chain gene variable domain contains 40x V domains, 25x D domains, 6x J domains and 5x C domains
How are the domains recombined in developing B-cells?
In developing B-cells these are recombined in a process known as VJ or V(D)J recombination
What is the process of VJ-recombination (light chain)?
- developing B cells join together separate gene segments in DNA in order to create the genes that encode the primary repertoire of low-affinity antibodies
- during the development of a B cell, a coding sequence joining a V to a J segment is assembled by removing the intervening genomic DNA (e.g. joining V3 to J3)
- transcription starts immediately upstream of the fused V segment, which lies immediately upstream of a J region. Extra downstream J segments are transcribed but edited out of the mRNA transcript
What is the process of V(D)J recombination (heavy chain)?
- The process of V(D)J recombination joins separate antibody gene segments together to form a functional VL- of VH- region coding sequence
- DNA splicing is driven by the V(D)J recombinase enzyme (encoded by RAG1 & RAG2 genes)
- RAG1 or RAG2 mutations have a severe combined immunodeficiency phenotype (SCIP). Such mice are often used in xenograft experiments
- during joining of gene segments, a variable number of nucleotides are often lost or inserted from the ends of the recombination gene segments. This is called JUNCTIONAL DIVERSIFICATION
- in many cases this will shift the reading frame to produce a nonfunctional gene. These developing B cells never make a functional antibody molecule and die in the bone marrow.
- Developing B&T-cells are diploid (one maternal & one paternal copy), but choose just one allele to recombine - this is known as allelic exclusion.
ALL OF THIS JUST GENERATES THE PRIMARY ANTIBODY REPERTOIRE.
How does the generation of mature antibodies?
- a naive, unchallenged human immune system contains around 1 x 10^12 different antibody molecules (the primary antibody repertoire
- a mature immune system can make antibodies able to bind to essentially antigen (essentially infinite flexibility)
Where do you get 1 x 10^12 antibodies from?
Antigen-driven somatic hypermutation
What occurs to the affinity of the antibodies after initial immunization?
Over time after initial immunization, there is a progressive increase in affinity of the antibodies towards antigens - this is known as affinity maturation.
What is the process of somatic hypermutation?
Accumulation of point mutations in both heavy and light chain V-region coding sequences. This happens AFTER VJ/V(D)J recombination has assembled the gene segments to generate the primary repertoire.
Go from low affinity to high affinity over time. Happens via feedback loop - cells that produce best antibodies proliferate the most.
What is somatic hypermutation?
- developing B-cells present their antibodies on their surface and binding of antigens stimulate their proliferation.
- most somatic mutations will either have no effect or will the antibody worse. This will stop antigen binding. Remove stimulus.
- cells containing mutations that increase affinity of the antibody to the antigen will increase the stimulus. These clones will survive and proliferate (especially as antigen levels get very low)
- in vivo evolution that selects cells with beneficial mutations
