Flashcards in Antibodies Deck (38)
What is the structure of immunoglobulin?
2 heavy chains and 2 light chains linked by disulphide bonds
They can be cleaved into 2 parts using protease papain into a Fab (fragment antigen binding) fragment and an Fc region (fragment crystallisation)
What are the FAB and Fc regions of an immunoglobulin?
The Fab fragment region is responsible for antigen binding (which there are 2 antigen binding domains per molecule)
The Fc region is bound by Fc receptors expressed by phagocytes and other immune cells
What are the two types of light chain?
What are the two types of domain?
Variable domain - made up of 2 domains: Variable light (VL) and Variable Heavy (VH)
They vary which defines antigen specificity
Constant domain - made up of 4 domains: Constant Light (CL) and 3 Contant Heavy (CH1, CH2 and CH3)
Within the same class they don't differ
What is the antigen binding site made up of?
The variable domain - Variable light (VL) and Varibale Heavy (VH)
There are loops that project form theses regions - called complementary determining regions (CDR) and hypervariable loops
The loops help determine specificty/complementarity
How were epitopes discovered?
Hapten - a small molcule that can act as an epitope but wouldn't induce antibodies on their own
1. Hapten is combined with a protein (ovalbumin)
2. Give the compound to a mouse
3. This will produce antibodies to ovalbumin and the hapten
4. Take the hapten antibodies and bind to hapten in vitro
What can act as an epitope?
Any chemical can act as an epitope (hapten) in this way; inorganic chemical, peptide, sugar, lipid, nucleic acid
Antibodies recognise antigenic epitopes by molecular complementation i.e. can't have steric interferences/hindrances
What is important about the epitope shape?
Variation in hapten structure (chemical positions of functional groups) can lead to different affinities of binding to an antibody
What binding do antibodies use?
Antibodies use non-covalent bonds to bind to antigenic epitopes:
Van-der Waals forces
A complementary binding surface on an antibody maximises the non-covalent interactions with the epitope
What are the types of epitopes?
Continuous epitopes - residues that make up the epitope are all directly linked
Discontinuous epitopes - formed by the apposition of distant residues as a result of molecular folding
Discontinuous epitopes are typically lost by unfolding or denaturing protein antigens
Epitopes can be 12 - 16 amino acids or 5 - 6 sugar residues
What are the 5 classes of immunoglobulins? What are they distinguished by?
IgM, IgG, IgA, IgE and IgD
They are distinguished by their heavy chains:
IgM - µ (mu) heavy chain
IgG - γ (gamma) heavy chain
IgA - a (alpha) heavy chain
IgE - ε (epsilon) heavy chain
IgD - δ (delta) heavy chain
Describe the structure of IgM?
Looks like a star
µ heavy chain has 4 CH domains
It is pentameric (970kDa): 10 antigen binding sites
The J chain promotes polymerisation
The monomers are linked by disulphide bonds to each other and to the J chain
Can be imaged by electron microscopy
What are the functions of IgM?
It is the first antibody secreted in response to a foreign antigen (primary response)
Agglutinates (clumps) microbes
Activates classical complement pathway
IgM is effective in limiting the spread of microorganisms via the bloodstream
Facts about IgM?
Confined to the vascular system due to large size
It has a low affinity for an antigen, but has 10 binding sites so can vind to polyvalent antigens e.g. Bacterial surfaces
Describe the structure of IgG?
Looks normal - monomeric
4 distinct isotypes: IgG1, IgG2, IgG3 and IgG4
IgG1, IgG2 and IgG4 - 146 kDa
IgG3 - 165 kDa
Each gamma-heavy chain has 3 CH domains
What are the functions of IgG?
Bound by Fc receptors on phagocytes
Activates the classical complement pathway
During pregnancy it is actively transported across placenta into the foetus to protect new born babies for the first 3-6 months
Facts about IgG?
Produced in secondary response to an antigen by B-cells in the lymph nodes and spleen
Major antibody in normal human blood, IgG1 is the most abundant isotype in the blood (9 mg/ml in serum)
Can enter the extra-vascular sites due to small size i.e. the tissues of the body
IgG opsonises microbes for uptake by phagocytes
Describe the structure of IgA?
Either normal - monomeric (160 kDa) or two joined together at the bottom - dimer (320 kDa)
In the dimer disulphide bonds and the J chain hold them together
What are the functions of IgA?
The principal function of IgA is to neutralise antigens:
IgA prevents bacterial toxins binding their cellular targets
IgA inhibits microbial adhesion to epithelia
IgA inhibits viral infectivity
Facts about IgA?
IgA is produced by B-cells in the mucosal associated lymphoid tissues during secondary immune response
Majority is secreted onto mucosal epithelial surfaces as Ig receptor transports it across the epithelium
Describe the structure of IgE?
Monomeric - 188 kDa
The ε heavy chain has 4CH domains
What are the functions of IgE?
Important for immune responses against helminths (worms) and other parasites
Also involved in type I hypersensitivity responses (allergic reactions) eg hay fever, food allergies
Binding of IgE molecules to polyvalent antigens cross-links Fc(epsilon) receptors and triggers release of inflammatory mediators
Facts about IgE?
Produced in response to some antigens, usually by B-cells in the mucosal associated lymphoid tissues• Low concentration in the blood
Vast majority of IgE is bound to Fc(epsilon) receptors on mast cells, basophils and eosinophils
Monomeric - 160 kDa
The δ heavy chain has 3CH domains
Acts as antigen receptor on immature B-cells and is present in the blood at very low concentrations
No other biological function has yet been described
What is antibody diversity? How does it arise?
Antibody diversity refers to the array of different antigenic specificities produced in the humoral immune response
Antibody diversity is generated by recombination of the antibody genes during the antigen-independent differentiation of B-cells in the bone marrow (antibody gene rearrangement)
Facts about antibody diversity?
5 X 10^13 different specificities of antibodies can be produced
This allows enough antibodies for every antigen it encounters
The total number of genes in the human genome is insufficient to accommodate one complete gene for each antibody molecule (there are only around 21,000 protein coding genes in the human genome)
Three of these genes are for immunoglobulin
Solution: Diversity is achieved by recombination of less than 1,000 gene segments in the DNA of developing B-cells
How does specificity arise?
Antigenic specificity of an antibody arises from the sequence of the VL and VH domains
In order to encode an antibody molecule, immunoglobulin genes must combine
What are all the gene segments that are used within the chains?
V coded for by variable (V)-gene segment [98aa]
D coded for by diversity (D)-gene segment [16-21aa]
J coded for by joining (J)-gene segment [1-15aa]
C coded for by constant gene segment
What is the Variable light chain coded for by? Example?
Variable (V) and Joining (J)
K-light chain of chromosome 2
39 V gene segments and 5 J gene segments
It also has a constant gene segment (C)