Lecture 5 - Antibody Structure, Function and Genetics Flashcards
Two types of antibodies
Membrane-bound and secreted
Light chain weight
25 kDa
Heavy chain weight
50kDa
Length of Ig domain
90-110 amino acids
Are there disulphide links between light chains?
No
Which parts make up antigen-binding domains?
Vh and Vl
Reason for flexible hinge region of antibody
Makes it less necessary for antigen to be placed very precisely relative to antibody binding site
CH2 domain role
Complement binding domain
CH3 domain role
Fc receptor binding site
Structure of Ig domain 1) 2) 3) 4) 5)
1) 90 - 100 amino acids long
2) 2 layers of beta-pleated sheet
3) between 3 - 5 antiparallel strands
4) Hydrophobic residues point inwards to form a hydrophobic core
5) Conserved cysteine regions form disulphide bridge
Classifications of Ig domains
V-like of C-like, depending on similarity to variable or constant regions
LIght chain constant region order of strands
DEBA - GFC
Heavy chain variable region order of strands
DEBA - GFCCC`
Which strands in a light chain B-pleated sheet join with a disulphide bridge?
B - F
What is the Ig superfamily?
Proteins that share structural and amino-acid homology with light or heavy chains
What can members of the Ig superfamily not be?
Cytosolic proteins
How are members of the Ig superfamily normally encoded?
As a single exon
Examples of members of the Ig superfamily 1) 2) 3) 4) 5) 6)
1) TCR
2) MHCI
3) MHC II
4) IgG
5) Cd4
6) CD8
Ig superfamily proteins are normally found where?
Bound in cell membranes
Two main proteases used to digest Ig domain
Pepsin
Papain
Where does papain cut?
Above disulphide hinge
Where does pepsin cut
Below disulphide hinge
Effects of cutting Ig domain with papain
2 regions:
1) Fab (can bind antigens)
2) Fc (can be crystallised)
Effects of cutting Ig domain with pepsin
1 large region:
Fab`2 - Can bind antigen, is dimeric
Other region degrades
Structure of antibody-binding site
6 loops:
3 from light chain
3 from heavy chain
Complementarity determining regions
3 loops of each chain that make up antibody-binding site
Another term for complementarity determining regions
Hypervariable loops
Location of hypervariable loops on V domain
At roughly 30 amino acids, 50 amino acids and 100 amino acids
Where are the hypervariable loops located in an antibody?
All exposed on the ends of a natively-folded protein
Ratio of kappa to lambda light chains in humans
~2 kappa:1 lambda
Are kappa and lambda light chains ever mixed in a single antibody?
No
Serum [IgG]
12g/L
Serum [IgA]
3g/L
Serum [IgM]
1.5g/L
Serum [IgD]
0.3g/L
Serum [IgE]
0.0005g/L
Why is serum [IgE] so low?
IgE is bound to the surfaces of cells, such as Mast cells
Therefore not present in the blood at high concentrations
Why is serum [IgD] so low?
Lacks molecular ability to be secreted.
Some IgD is present in the blood because it can be cleaved off the surfaces of cells
Function of IgD
Cell surface receptor
Number of IgM mu chains
10
Does membrane-bound IgM have a J chain?
No
IgG distribution
Serum and lymph
IgM distribution
Serum
IgA distribution
Serum and mucosal secretions
IgE distribution
Below epithelial surfaces
IgG features 1) 2) 3) 4)
1) Monomer
2) Major Ig in secondary response
3) Large, flexible hinge region
4) High affinity
IgM features 1) 2) 3) 4)
1) Pentamer
2) Has extra C domain (3 C domains, 3 of which make up Fc tail structure)
3) High avidity, low affinity
4) 26aa ‘tail piece’ linked to J chain
J chain
15kDa protein involved in Ig oligeromisation
Bound to IgM and IgA
Affinity
The strength of binding between two molecules at a single site
Avidity
Sum total of strength of binding between two molecules, where multiple binding-sites are involved
IgA features 1) 2) 3) 4)
1) Dimer (tetravalent)
2) Major Ig secreted in GIT and respiratory tracts
3) 26aa tailpiece bound to J chain
4) Uses secretory component to bind to pIgR to transcytose across luminal membrane into luminal space
IgE features 1) 2) 3) 4)
1) Has extra Fc domain
2) Involved in allergic reactions and anti-parasitic immunity
3) Inflexible hinge region
4) Binds to mast cells, induces degranulation
