Lecture 11 Flashcards
What are antibodies?
Soluble globular proteins produced by B lymphocytes and secreted into the serum, lymph or at mucosal surfaces
These are a key component of the adaptive immune response in higher vertebrates with antibodies conferring resistance to pathogens through recognition of an antigenic epitope on the surface of a pathogen
Where are antibodies found?
They are found in the gamma globulin fraction of serum and a normal healthy individual will have about 20-30mg/ml or serum immunoglobulin making them the second most abundant serum proteins after albumin
What is the Ig domain?
This a basic and highly successful protein fold of which antibodies are repeats of, however this fold is also found in many other proteins such as T cell receptors
What experiment was first performed by Elvin Kabat and George Wu?
These scientists were the first to determine structural information about the antibody where they worked with a patient in late stage multiple myeloma
From there work they determined that there were 3 highly conserved regions of the structure termed framework regions and 3 regions of extreme amino acid variability termed hypervariable regions
There were also always two completely conserved cysteine residues located in the FR1 and FR3 regions which formed the intra-chain disulphide bond that give Ig domain great stability
Where were the hyper-variable regions initially found in the Ig domain?
HV1 is between amino acids 25-35
HV2 is between amino acids 55-65
HV3 is between amino acids 95-110
What are the features of the beta barrel Ig structure?
It has two antiparallel beta sheets made up of seven (constant) or nine (variable) b-strands connected by unconstrained loops
These two sheets lie parallel but at a 30 degree twist to form a very soluble beta barrel structure which is further stabilized by a single disulphide bond
The unconstrained loops is what allows the extreme amino acid diversity without compromising the integrity of the structure
What are the features of the light chain of the Ig structure?
This is 25 kD in size composed of one variable (VL) and one constant (CL) domain with the amino acids in the VL varying considerably between different Ig molecules while those in the C region amino acid sequences are invariant between antibodies
These gene are at two loci with the lambda and kappa being equally used in humans
What are the features of the heavy chain of the Ig structure?
These are 50-75 kD consisting of one variable domain with 3-5 constant Ig domains
These changes have a hinge region where heavy chains join to each other where there are 1-7 disulphide bonds depending on which class (IgG, IgM, IgA, IgD and IgE)
What does the antibody molecule consist of?
There are 2 heavy chains and 2 light chains with the H chains being joined at the hinge region via 1-7 disulphide bonds while the L chains are joined to the H chains by an inter chain disulphide
This makes the general formula for antibodies H2L2 though IgA and IgM exist in dimer and pentameric form respectively
The total molecular weight of an IgG molecule is 150 kD
What experiments did Edelmen and Porter do, and what did they determine about the Ig structure?
They made use papain to cleave IgG molecules, this cleaved it into two parts above the hinge region with one part retaining the ability to bind antigen being termed the Fab fragment while the other part crystalized very easily and retained the ability to block complement and was termed the Fc fragment
They also used pepsin which could produce a much larger protein fragment which contained the antigen binding activity whch resolved into 2 50 kD Fab fragments on mild reduction which were called the Fab2’ fragment
From this they deduced that papain cleaved once above the disulphide and pepsin cleaved once below the disulphide and then multiple times I the Fc region allowing them to predict the famous Y shaoed molecule of IgG
What are the functions of IgG?
major serum isotype placental immunity soluble anti-toxin anti-bacterial surface opsonises bacteria initiates or “fixes” complement blocks receptor binding high affinity and specificity there are 4 subclasses in humans but only 2 in mice with 2 isotypes
What are the functions of IgM?
Membrane bound and soluble forms
mIgM is the default B cell receptor
consist of 5 chains joined by an invariant J-chain
IgM is the default isotype produced by all naïve B cells
This invariably has low affinity but avidity due to its ability to form pentamers
It is surface reactive and binds with high avidity
It is an opsonin
Fixes complement
First line of serum defense
What are the different H chain genes for the Ig classes?
IgG uses the gamma gene IgA uses the alpha gene IgM uses the mew gene IgD uses the delta gene IgE uses the epsilon gene
What are the functions of IgA?
Produced in serum and as dimer at mucosal surfaces such as the gut, lungs, saliva and breast milk
Secreted form is linked by J chain and secretory component (protection and transport across the epithelium)
Primary protection against pathogens at mucosal surfaces
First line of defense at mucosal surface
Blocks pathogen adhesion and invasion
What are the functions of IgD?
This has a membrane and a soluble form however the role of the soluble form is not known
B cells with mIgD but not mIgM ressit central tolerance
