Antibody

Structure & Functions

Antibody structure key features?
Antibody structure
H H
IgA a chains
IgE e
IgM m
IgG g
IgD d
k, l
2 Heavy chains- different for each class
2 Light chains- k or l for all classes
COOH COOH
NH3
NH3
NH3
NH3
Polypeptide chains or glycopeptide (decorated with CHO)
Held together with disulphide bonds
Antibody fragments
H H
H H
Pepsin
F(ab)2
Fab
H H
Papain
Fc
Detect antigen
Precipitate antigen
Block the active sites of toxins or pathogen-associated
molecules
Block interactions between host and pathogen-associated
molecules
(Fab)
2
fragment
Inflammatory and effector functions associated with cells
Inflammatory and effector functions of complement
The trafficking of antigens into the antigen processing
pathways
Fc fragment
Antibody is organised into domains
antigen
antigen
Variable
region (V)
Constant
region (C)
Light chain (
L
)
21-22KDa
Heavy chain (
H
)
52-64KDa
V region
Controls antigen
specificity
Fc region
Controls
effector
function
m and e have 4 CH domains and no hinge region
Ab usually have 3CH domains
A domain is the characteristic structural motif of all Ig domains
is a b barrel of 7 (CL) or 8 (VL) polypeptide strands
connected by loops
arranged to enclose a hydrophobic interior
eg. Single VL domain
Hinge
CH3
CH2
CH1
VH1
VL
CL
Elbow
Crystal structure showing antibody domains
Structurally antibodies are arranged into domains.
Domains are derived from a single ancestral gene that
has duplicated, diversified & modified.
Ig-like Domains are not restricted to immunoglobulins.
The characteristic of the immunoglobulin domain is a
disulphide bond (110aa)
Immunoglobulin domains
The genes encoding Ig domains
are not restricted to Ig genes.
They were subsequently found
in a superfamily of related genes
They encode proteins for cell-
cell interactions and molecular
recognition.
Found in most cell types and
different animal species
Ig gene superfamily
Immunoglobulins as Bifunctional Proteins
Immunoglobulins must interact with a small number of
specialised molecules (common Fc to each class)
Fc receptors on cells
Complement proteins
Intracellular cell signalling molecules

whilst simultaneously recognising an infinite array of
antigenic determinants (? Mechanism).
FR1 FR2 FR3 FR4 CDR2 CDR3 CDR1
Distinct regions of high variability and conservation led to the concept
of a framework on which hypervariable regions were suspended.
Framework and Hypervariable regions
Amino acid No.
Variability
80
100
60
40
20
20 40 60 80 100 120
Most hypervariable regions coincide with antigen contact points -
the COMPLEMENTARITY DETERMINING REGIONS (CDRs)
The sequences of the hyper-variable loops :-
are highly variable amongst antibodies of different
specificities
influence the shape, hydrophobicity and charge at the
tip of the antibody
accounts for the diversity of antigens that can be
recognised
Hypervariable loops
L & H chain folds to yield 3 CDR in each chain
to form walls of Ag binding groove
Hypervariable regions
Hypervariable CDRs are located on loops at the end of the
Fv regions
Activates C1q complement
(when bound to Ag)

Opsonises (by binding
to phagocyte and Ag)
Helps kill infected cells
(eg.via ADCC,
eg. phagocytosed organisms)
Activates immune cells
(by binding to cell and Ag)
Summary of antibody effector
functions?
IgM IgG
IgD IgM - B cell receptor
IgE binds to mast cells
IgG- macrophages
IgG - blood
IgA - mucosa
Mainly IgG
EFFECTOR FUNCTIONS of antigen-bound antibody
Effector function is determined by Antibody class or
subclass How? This is largely determined by the
distribution of specific Fc receptors for each class of Ab
Fcg HIGH AFFINITY Fc receptors
Receptor Cell type Effect of ligation
FcgRI Macrophages Neutrophils,
Eosinophils, Dendritic cells Uptake, Respiratory burst
FcgRIIA Macrophages, Neutrophils,
Eosinophils, Platelets
Langerhans cells Uptake, Granule release
FcgRIIB1 B cells, Mast Cells No Uptake, Inhibition of stimulation
FcgRIIB2 Macrophages ,Neutrophils,
Eosinophils Uptake, Inhibition of stimulation
FcgRIII NK cells, Eosinophils,
Macrophages, Neutrophils,
Mast cells Induction of killing (NK cells)
Monomeric IgM
IgM only exists as a monomer on the surface of B cells
Cm4 contains the transmembrane and cytoplasmic regions. These
are removed by RNA splicing to produce secreted IgM
Monomeric IgM has a very low affinity for antigen
Cm2
N.B. Only constant
heavy chain
domains are shown
Cm1 binds C3b to facilitate uptake of opsonised antigens by
macrophages
Cm4 mediates multimerisation (Cm3 may also be involved)
Cm3 binds C1q to initiate activation of the classical
complement pathway
Polymeric IgM
IgM forms pentamers and hexamers
Non-antigen binding sites
IgM facts and figures
Heavy chain: m - Mu
Half-life: 5 to 10 days
% of Ig in serum: 10
Serum level (mgml
-1
): 0.25 - 3.1
Complement activation: ++++ by classical pathway
Interactions with cells: Phagocytes via C3b receptors
Epithelial cells via polymeric Ig receptor
Transplacental transfer: No
Affinity for antigen: Monomeric IgM - low affinity - valency of 2
Pentameric IgM - high avidity - valency of 10
IgD facts and figures
IgD is co-expressed with IgM on B cells due to differential RNA splicing
IgM > IgD on nave B cells
function of IgD in host defence is unknown - knockout mice inconclusive
Ligation of IgD with antigen can activate, delete or anergise B cells
Heavy chain: d - Delta
Half-life: 2 to 8 days
% of Ig in serum: 0.2
Serum level (mgml
-1
): 0.03 - 0.4
Complement activation: No
Interactions with cells: T cells via lectin like IgD receptor
Transplacental transfer: No
IgA dimerisation and secretion
IgA is the major isotype of antibody secreted
at mucosal surfaces

Exists in serum as a monomer, but also as a
J chain-linked dimer
IgA1 is mostly found in serum (monomer), made by
bone marrow B cells

IgA2 is mostly found in mucosal secretions (dimer),
colostrum and milk; made by mucosal B cells
Body eg gut or nasal mucosa
Mucosal
epithelium
1. IgA2 binds to
pIgR
2. Bound IgA
is transported
across epithelium
3. sIgA
is released via
cleavage of pIgR
IgA facts and figures
Heavy chains: a1 or a2 - Alpha 1 or 2
Half-life: IgA1 5 - 7 days
IgA2 4 - 6 days
Serum levels (mgml
-1
): IgA1 1.4 - 4.2
IgA2 0.2 - 0.5
% of Ig in serum: IgA1 11 - 14
IgA2 1 - 4
Complement activation: IgA1 - by alternative and lectin pathway
IgA2 - No
Interactions with cells: Epithelial cells by pIgR
Phagocytes by IgA receptor
Transplacental transfer: No
IgA is inefficient at causing inflammation and elicits protection by excluding, binding,
cross-linking microorganisms and facilitating phagocytosis
IgE facts and figures
IgE has a role in protecting against parasite infections
Most IgE is absorbed onto the high affinity IgE receptors of effector cells
IgE is also closely linked with allergic diseases
Heavy chain: e - Epsilon
Half-life: 1 - 5 days
Serum level (mgml
-1
): 0.0001 - 0.0002
% of Ig in serum: 0.004
Complement activation: No
Interactions with cells: Via high affinity IgE receptors expressed
by mast cells, eosinophils, basophils
and Langerhans cells
Via low affinity IgE receptor on B cells
and monocytes
Transplacental transfer: No
The high affinity IgE receptor (FceRI)
a chain
b chain
g2
S S
S S
S S
The IgE - FceRI interaction is
the highest affinity of any Fc
receptor with an extremely low
dissociation rate.

CH3 of IgE interacts with the
FceRI causing a conformational
change.

Binding of IgE to FceRI thus
increases the half life of IgE

IgG facts and figures
Heavy chains: g 1 g 2 g3 g4 - Gamma 1 - 4
Half-life: IgG1 21 - 24 days IgG2 21 - 24 days
IgG3 7 - 8 days IgG4 21 - 24 days
Serum level (mgml
-1
): IgG1 5 - 12 IgG2 2 - 6
IgG3 0.5 - 1 IgG4 0.2 - 1
% of Ig in serum: IgG1 45 - 53 IgG2 11 - 15
IgG3 3 - 6 IgG4 1 - 4
Complement activation: IgG1 +++ IgG2 +
IgG3 ++++ IgG4 No
Interactions with cells: All subclasses via IgG receptors on macrophages
and phagocytes
Transplacental transfer: IgG1 ++ IgG2 +
IgG3 ++ IgG4 ++
Carbohydrate is also
essential for complement
activation
C1q binding motif on
the Cg2 domain

Subtle differences in hinge region between subclasses accounts for
differing abilities to activate complement
a, m, d, e, g
k, l chains
Species specific C (Hyper)-variable region
(antigen binding site)
ANTIBODY variability
isotypic allotypic idiotypic