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HKMU
• Faculty of Medicine
Department of biochemistry & Molecular
Biology
• First semester January 2016
• BASIC IMMUNOGLOBULIN STRUCTURE
Immunoglobulins generally assume one of
two roles: immunoglobulins may act as
• plasma membrane bound antigen receptors
on the surface of a B-cell or ii) as antibodies
free in cellular fluids functioning to intercept
and eliminate antigenic determinants.
• In either role, antibody function is intimately
related to its structure
• Immunoglobulins:
• Are composed of four polypeptide chains:
two "light" chains (lambda or kappa), and two
"heavy" chains
• 1.alpha 2.delta 3. gamma 4.epsilon 5.mu
• The type of heavy chain determines the
immunoglobulin isotype (IgA, IgD, IgG, IgE,
IgM, respectively).
• Light chains are composed of 220
amino acid residues while heavy chains
are composed of 440-550 amino acids.
• -Each chain has "constant" and
"variable" regions.
• Variable regions are contained within
the amino (NH2) terminal end of the
polypeptide chain (amino acids 1-110).
• When comparing one antibody to another,
these amino acid sequences are quite
distinct.
• Constant regions, comprising amino acids
111-220 (or 440-550), are rather uniform, in
comparison, from one antibody to another,
within the same isotype.
• "Hypervariable" regions, or
"Complementarity Determining Regions"
(CDRs) are found within the variable regions
of both the heavy and light chains.
• These regions serve to recognize and
bind specifically to antigen. The four
polypeptide chains are held together by
covalent disulfide (-S-S-) bonds
Structural differences between
immunoglobulins are used for their
classification
-
-. As stated above, the type of heavy chain
an immunoglobulin possesses determines
the immunoglobulin "ISOTYPE".
• More specifically, an isotype is determined by
the primary sequence of amino acids in the
constant region of the heavy chain, which in
turn determines the three-dimensional
structure of the molecule.
• Since immunoglobulins are proteins, they
can act as an antigen, eliciting an immune
response that generates anti-immunoglobulin
antibodies.
• However, the structural (three-dimensional)
features that define isotypes are not
immunogenic in an animal of the same
species, since they are not seen as "foreign".
• For example, the five human isotypes, IgA,
IgD, IgG, IgE and IgM are found in all humans
and a result, injection of human IgG into
another human would not generate antibodies
directed against the structural features
(determinants) that define the IgG isotype
• . However, injection of human IgG into a
rabbit would generate antibodies directed
against those same structural features.
• Another means of classifying
immunoglobulins is defined by the term
"allotype". Like isotypes, allotypes are
determined by the amino acid sequence and
corresponding three-dimensional structure of
the constant region of the immunoglobulin
molecule
• Unlike isotypes, allotypes reflect genetic
differences between members of the same
species.
• This means that not all members of the
species will possess any particular allotype.
Therefore, injection of any specific human
allotype into another human could possibly
generate antibodies directed against the
structural features that define that particular
allotypic variation
• A third means of classifying immunoglobulins
is defined by the term "idiotype". Unlike
isotypes and allotypes, idiotypes are
determined by the amino acid sequence and
corresponding three-dimensional structure of
the variable region of the immunoglobulin
molecule. In this regard, idiotypes reflect the
antigen binding specificity of any particular
antibody molecule
• BASIC IMMUNOGLOBULIN
FUNCTION
• Antibodies function in a variety of ways
designed to eliminate the antigen that
elicited their production. Some of these
functions are independent of the
particular class (isotype) of
immunoglobulin
• These functions reflect the antigen binding
capacity of the molecule as defined by the
variable and hypervariable (idiotypic) regions.
• For example, an antibody might bind to a
toxin and prevent that toxin from entering host
cells where its biological effects would be
activated
• Similarly, a different antibody might bind
to the surface of a virus and prevent
that virus from entering its host cell. In
contrast, other antibody functions are
dependent upon the immunoglobulin
class (isotype). These functions are
contained within the constant regions of
the molecule.
• For example, only IgG and IgM antibodies
have the ability to interact with and initiate the
complement cascade.
• Likewise, only IgG molecules can bind to the
surface of macrophages via Fc receptors to
promote and enhance phagocytosis.
GENERATION OF
ANTIBODY DIVERSITY
• The immune system has the capacity to
recognize and respond to about (Ten million)
107 different antigens.
• This extreme diversity can be generated in at
least three possible ways:
1. Multiple genes in the germ line DNA.
2. Variable recombination during the
differentiation of germ line cells into B-cells.
3. Mutation during the differentiation of germ line
cells into B-cells.
The figure shows the genetic makeup of a germ line cell and a
mature B-cell at the loci controlling heavy chain production.
Germ line DNA has many (up to 200) different variable (V)
region genes, in addition to 12 diversity (D) region genes and
four joining (J) region genes.
• During differentiation of this cell into the Bcell, rearrangement of the DNA occurs.
• This rearrangement aligns one of the
many V genes with one of the D genes
and one of the J genes, producing a
functional VDJ recombinant gene.
• Since any of the genes may recombine
with any others, this rearrangement has
the potential to generate 200(v) x 12(D) x
4(J) = 9600 different possible
combinations
1. The same type of event occurs in the
genes encoding the immmunoglobulin
light chains where about 200 different V
regions may recombine with about 5
different J regions giving rise to 200 x 5 =
1000 possible light chains.
1. Since in any particular B-cell, any light
chain combination can occur along with
any heavy chain combination, the total
possible immunoglobulin combinations
approaches 107 (9600 x 1000).
• Antibodies:
• Are glycoproteins
• are built of subunits containing
– two identical light chains (L chains), each containing about 200 amino
acids
– two identical heavy chains (H chains), which are at least twice as long
as L chains
• The first 100 or so amino acids at the N-terminus of both H
and L chains vary greatly from antibody to antibody – they are
termed variable (V) regions
– unless members of the same clone (often not even then!), no two B
cells are likely to secrete antibodies with the same V region
– the amino acid sequence variability in the V regions is especially
pronounced in 3 hypervariable regions also called CDR’s
– the tertiary and quaternary structure of the antibodies brings the 3
CDR’s of both the H and L chains together to form the antigen
binding site which binds the antigen epitope
• Only a few different amino acids sequences are found in the Cterminus of the H and L chains and these are called constant (C)
regions
– two different kinds of C regions for L chains -- kappa (κ) and
lambda (λ)
– five different kinds of C regions for H chains
• mu (µ) chains, IgM
• gamma (γ) chains, IgG
• alpha (α) chains, IgA
• epsilon (ε) chains, IgE
• delta (δ) chains, IgD
– each of these 5 kinds of H chains can pair with either lambda or
kappa L chains
Constant-Region Domains
• The carboxyl-terminal domains of immunoglobulins display considerably
less sequence variability within a given isotype than observed for V-region
domains. These domains are referred to as constant (C) regions.
• H-chain C regions are numbered (CH1, CH2, CH3, and CH4) beginning
with the most V-region proximal domain.
• The C region domains of the H-chain have been shown to be responsible
for many aspects of antibody function, including interaction with Fc
receptors, complement fixation, transplacental transfer, the ability to form
multimers, and the capacity to be secreted across mucosal surfaces.
• Because different H-chain isotypes have different C region domains, these
capabilities vary with the class of the particular antibody.
• Five major classes of H-chain C regions exist: (γ, α, µ, δ, ε). As a direct
consequence of the correlation between the H-chain class of an antibody
and its resultant effector functions, immunoglobulins have been named
according to there heavy chain, i.e., IgG, IgA, IgM, IgD and IgE.
Hinge Region
• Immunoglobulins have a hinge region located C-terminal of the the CH1
domain of their H-chains.
• In the case of the H-chains of the µ and ε isotypes the hinge is so elongated
that it is actually an extra immunoglobulin domain explaining the presence
of a fourth C domain in these isotypes.
• Other heavy chain isotypes (γ, α, and δ) use shorter stretches of
polypeptide, which are thought nonetheless to have evolved from the Cµ/ε2
domain.
• Hinge regions are encoded by separate exons.
• The hinge region permits a considerable degree of flexibility between the
antigen-binding and effector-interacting components of the
immunoglobulin molecule.
Concept: All classes of antibody have the same basic structure.
Two identical heavy chains and two identical light chains: each of
which contributes to antigen binding.
The Structural Organization of the Human Immunoglobulin Isotypes.
Structure of the 5 Major Classes of Antibody
Immunoglobulin G (IgG)
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The most abundant class in serum ~80% of the total serum immunoglobulins.
IgG is found also in the interstitial spaces.
The IgG molecule consists of two λ H-chains and two κ or two λ L-chains.
In man there are four subclasses of IgG.
IgG fixes complement
IgG is the only immunoglobulin to cross the placenta.
IgG reacts with FcR’s on phagocytic cells to promote opsonization.
IgD and IgE
• The principal role of IgD: Located on the surface of B-lymphocytes
and along with IgM serves the function of antigen recognition by the B
cell.
• IgE binds to mast cells and basophils through a high affinity FcR and
is involved in mediating hypersensitivity reactions.
Study questions
1. Structures of all immunoglobulins
2. Molecular basis of ab diversity
3.Terminologies ie allotypes,
idiotypes etc