ANTIGEN RECOGNITION IN THE

ADAPTIVE IMMUNE SYSTEM

Sitti Wahyuni, MD, PhD

Department of Parasitology,
Medical Faculty, Hasanuddin University

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 DURING LECTURE/DISCUSSION

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• Function of cellular receptors is to detect external stimuli and
trigger responses of the cells on which the receptors are
expressed.
• Antigen receptors of lymphocytes must be able to bind to and
distinguish between many, often closely related, chemical
structures.
• Antigen receptors on lymphocyte B and T are clonally distributed
• The total number (repertoire) of lymphocyte specificities is very
large
• Antigen receptors transmit biochemical signals that are
fundamentally the same in all lymphocytes and are unrelated to
specificity
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 Questions to be addressed
• How do the antigen receptors of lymphocytes recognize
extremely diverse antigens and transmit quite conserved
activating signals to the cells?
• How is the vast diversity of receptor structures generated in
lymphocytes?

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 Antigen Receptors of Lymphocytes
• Consist:
– variable (V) regions
– constant (C) regions.
• In V regions there are
variability called
hypervariable regions/
complementarity-
determining regions
(CDRs)

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 T cell receptors (TCR)
• Consist of α and β chain
• Each chain consist of
– Constant region (attach to T
cell membrane)
– Variable region (recognize
MHC molecules & Peptides
• Note
– Each TCR recognizes 1-3 peptide
of antigen
– Only a few peptides antigen (the
immunodominant epitopes), are
actually recognized by TCR

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 T cell receptors (TCR)
• The V region of each TCR chain
has 3 hypervariable/
complementarity-determining
(CDR).
• The CDR3 is the most variable
among different TCRs.

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 B cell receptors (BCR)
• Is the antibody M or D (called
membrane antibody)
• Composed of 4 chain
– 2 light chains
– 2 heavy chain
• Each chain containing
– variable (V) region to recognize
antigen
– constant (C) region anchored in
the B plasma membrane

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 TCR/ BCR Complex

• Antigen receptors are noncovalently attached to other

molecules to deliver signal to the inside of the cell
• Complex antigen receptors-signaling molecules
– in B lymphocytes: B cell receptor (BCR) complex,
– in T lymphocytes: T cell receptor (TCR) complex.
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• γδ TCR
– Lymphocyte T in innate immunity
– 5% - 10% of total T cells in the body
– Structurally similar to the αβ TCR but have very
different specificities
– May recognize a variety of protein and non-
protein antigens that are not displayed by MHC
molecules.
– Abundant in epithelia.

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• NK-T cells
– Less than 5% of all T cells
– Express markers of natural killer (NK) cells and are
called NK-T cells.
– NK-T cells
• Express αβ TCRs
• Recognize lipid antigens displayed by nonpolymorphic
class I MHC-like molecules
– The functions are not well understood.

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 Soluble and membrane Ig

• Soluble Ig • Membrane
– Recognize antigen – Recognize antigen
– Effector of lymphocite B – Consist of IgM and IgD
– Consist of IgG, IgA, IgM,
IgD, IgE
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 Antibodies

Antibody molecule
• Composed by 4 polypeptide
chains= 2 identical heavy (H)
chains & 2 identical light (L) chains
• Each chain containing variable
region & constant region
• The four chains are assembled to
form a Y-shaped molecule.
• Each light chain is attached to one
heavy chain

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 Antibodies

Antibody molecule
• 2 heavy chains are attached to
each other by disulfide bonds.
• The light chain is made up of
one V and one C domain
• The heavy chain has one V and
three or four C domains.

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• Variable region
– Regio that recognize & bind to
antigen
– Heavy chain called VH and light
chain called VL
– Contains 3 hypervariable regions
(CDRs), the greatest variability is
in CDR3, which is located at the
junction of the V and C regions
• Constant region
– Region that is not attached to
antigen
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• Regions of antibody based on the
properties of proteolytic fragments of
immunoglobulins.
– Fab (fragment antigen binding)
– Fc (fragment cystalline)
• Hinge region
– Between the Fab and Fc regions
– A flexible portion called the hinge
region & allows the 2 antigen-binding
Fab regions of each antibody
molecule to move independently of
each other

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• Light chain
– 2 types: κ and λ
– κ and λ, differ in their C regions, do not differ in function.
– Each B cell expresses either κ or λ, but not both
• Heavy chain
– 5 types: μ, δ, γ, Ε, and α (differ in their C regions)
– Antibodies that contain different heavy chains are said to
belong to different isotypes, or classes, and are named
according to their heavy chains (i.e., IgM, IgD, IgG, IgE, and
IgA)

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 Features of the major isotypes (classes) of antibodies
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• The antigen-binding regions of antibody
– Capable of binding a wide variety of antigens,
– Capable of accommodating many different shapes
– Antibodies bind to antigens by reversible,
noncovalent interactions, including hydrogen
bonds and charge interactions.
• The parts of antigens that are recognized by
antibodies are called epitopes, or determinants.

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• Affinity
– The strength of one antigen-binding surface of an
antibody to binds to one epitope of an antigen
– Is expressed as the dissociation constant (Kd)
– Kd is the molar concentration of an antigen required
to occupy half the available antibody molecules in a
solution
– The lower the Kd, the higher the affinity.
– Primary immune response: the Kd 10-6 to 10-9 M
– Secondary immune response: the Kd 10-8 to 10-11 M
• Affinity maturation: the increase in antigen-binding
strength
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• Antigen-binding site
– The site where antibody bind to epitope
– IgG, IgD, and IgE antibody molecule has two antigen-
binding sites
– IgA has 4 antigen-binding sites
– IgM has 10 antigen-binding sites
• Avidity
– The total strength of binding by
– Much greater than the affinity.
• Antibodies produced against one antigen may bind other,
structurally similar, antigens cross-reaction.
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• Monoclonal antibodies
– Antibody that arise from single clone of B cells that
only recognize single epitope of antigen
– Has far-reaching implications for clinical medicine
and research (diagnosis and therapy)

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• Most of antibody monoclonal are made by fusing cells from
immunized mice with mouse myelomas
• These mouse monoclonal antibodies cannot be injected
repeatedly into human subjects it will be regarded by
immune system as foreign
• Solution: the antigen-binding V regions is retained & the rest
is replaced with human Ig "humanized" antibody

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 Development of Immune Repertoires

• How the enormous diversity of these B and T cels receptors is

produced?
• There are many clones of lymphocytes with distinct
specificities, perhaps as many as 109, and these clones arise
before encounter with antigen.
• There are not enough genes in the human genome for every
possible receptor to be encoded by a different gene.
• The immune system has developed mechanisms for
generating extremely diverse repertoires of B and T
lymphocytes
• The generation of diverse receptors is intimately linked to the
process of lymphocyte maturation.
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 Maturation of lymphocytes

• The maturation of lymphocytes from bone marrow

stem cells consists of three types of processes:
– proliferation of immature cells
– expression of antigen receptor genes
– selection of lymphocytes that express useful
antigen receptors .
• Involving random genetic recombination events

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Interleukin-7 (IL-7)
• Produced by stromal cells in the bone marrow and
the thymus.
• Function:
– Stimulate proliferation of lymphocyte
– Maintains and expands the number of lymphocyte
progenitors

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 Steps in the maturation of lymphocytes
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• Antigen receptors are encoded by several gene
segments that are separate from one another in the
germline and recombine during lymphocyte
maturation.
• Diversity is generated during this recombination
process mainly by varying the nucleotide sequences
at the site of recombination.
• The expression of diverse antigen receptors is the
central event in lymphocyte maturation

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• Maturing lymphocytes are selected at multiple steps
during their maturation to preserve the useful
specificities.
• Selection is based on the expression of intact antigen
receptor components and what they recognize.
– Positive selection: Only select lymphocytes that
express antigen receptor
– Negative selection: Only select lymphocytes that
has low affinity to self antigens present in the
bone marrow and thymus
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 Production of diverse antigen receptors

• The expression of B and T lymphocyte antigen receptors is

initiated by somatic recombination of gene segments that
code for the variable regions of the receptors, and diversity is
generated during this process.
• Hematopoietic stem cells in the bone marrow as well as early
lymphoid progenitors contain Ig and TCR genes in their
inherited, or germline, configuration.

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• Each loci of heavy & light chain of BCR and α & β chain of TCR
contain
– Multiple variable region (V) genes
– One or a few constant region (C) genes
– Several small stretches of nucleotides, called joining (J)
– Diversity (D) gene segments.
• All antigen receptor gene loci contain V, J, and C genes
• The Ig heavy chain and TCR β loci also contain D gene segments
• Gene recombination is combination V, J, and C segment genes
(with or without D) the segments are selected randomly

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Recombination andS.Wahyuni/lecture/immunology/S1
expression of immunoglobulin (Ig) genes 37
 VDJ recombinase enzyme
• Mediate the somatic recombination of V-J, or V-D-J gene segments
• Its lymphoid-specific component of the VDJ recombinase
(recombinase-activating gene (RAG)-1 and RAG-2 proteins)
recognizes DNA sequences that flank all antigen receptor V, D,
and J gene segments.
• The enzyme brings the V, D, and J segments close together and
cleaves the DNA at specific sites.
• The DNA breaks are then repaired by ligases, producing a full-
length recombined V-J or V-D-J gene without the intervening DNA
segments
• The lymphoid-specific component of the VDJ recombinase is
expressed only in immature B and T lymphocytes.

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• Diversity of antigen receptors is produced by:
– the use of different combinations of V, D, and J gene
segments in different clones of lymphocytes (called
combinatorial diversity)
– by changes in nucleotide sequences introduced at the
junctions of V, D, and J gene segments (called junctional
diversity)
• Combinatorial diversity is limited by the number of available V,
D, and J gene segments, but junctional diversity is almost
unlimited.
• This junctional diversity is produced by three types of sequence
changes, each of which generates more sequences than are
present in the germline
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genes.
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 Mechanisms of diversity in antigen receptors
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Maturation and selection of
B lymphocytes

Steps in the maturation and selection of

B lymphocytes (Occur in Bone marrow)

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Maturation and selection of
T lymphocytes
• Largely related to the specificity of different subsets of T cells
for peptides displayed by different classes of MHC molecules.
• The most immature progenitors are called pro-T cells or
double-negative T cells (do not express CD4 or CD8).
• These cells expand in number mainly under the influence of
IL-7 produced in the thymus.
• Some of the progeny of double-negative cells undergo TCR β
gene recombination, mediated by the V(D)J recombinase.

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 Steps in the maturation and selection of major
2/12/2018 histocompatibility S.Wahyuni/lecture/immunology/S1
complex (MHC)-restricted T lymphocytes 44
 SUMMARY
• In the adaptive immune system, the molecules responsible for
specific recognition of antigens are antibodies and T cell
antigen receptors.
• Antibodies (also called immunoglobulins) may be produced as
membrane receptors of B lymphocytes and as proteins
secreted by antigen-stimulated B cells that have differentiated
into antibody-secreting plasma cells. Secreted antibodies are
the effector molecules of humoral immunity, capable of
neutralizing microbes and microbial toxins and eliminating
them by activating various effector mechanisms.
• TCRs are membrane receptors and are not secreted.

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• The core structure of antibodies consists of two heavy chains
and two light chains forming a disulfide-linked complex. Each
chain consists of a variable (V) region, which is the portion
that recognizes antigen, and a constant (C) region, which
provides structural stability and, in heavy chains, performs the
effector functions of antibodies.
• T cell receptors consist of an α chain and a β chain. Each chain
contains one V region and one C region, and both chains
participate in the recognition of antigens, which for most T
cells are peptides displayed by MHC molecules.
• The V regions of Ig and TCR molecules contain hypervariable
segments, also called complementarity-determining regions,
which are the regionsS.Wahyuni/lecture/immunology/S1
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of contact with antigens. 46
• The genes that encode antigen receptors consist of multiple segments that
are separate in the germline and are brought together during the
maturation of lymphocytes. In B cells, the Ig gene segments undergo
recombination as the cells mature in the bone marrow, and in T cells the
TCR gene segments undergo recombination during maturation in the
thymus.
• Receptors of different specificities are generated in part by different
combinations of V, D, and J gene segments. The process of recombination
introduces variability in the nucleotide sequences at the sites of
recombination by adding or removing nucleotides from the junctions. The
result of this introduced variability is the development of a diverse
repertoire of lymphocytes, in which clones of cells with different antigen
specificities express receptors that differ in sequence and recognition, and
most of the differences are concentrated at the regions of gene
recombination.
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• During their maturation, lymphocytes undergo alternating cycles of
proliferation and antigen receptor expression and traverse several
checkpoints at which they are selected such that only cells with complete
functional antigen receptors are preserved and expanded. In addition, T
lymphocytes are positively selected to recognize peptide antigens
displayed by self MHC molecules.
• Immature lymphocytes that strongly recognize self antigens are negatively
selected and prevented from completing their maturation, thus
eliminating cells with the potential of reacting in harmful ways against self
tissues.

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 Review questions

• What are the functionally distinct domains (regions) of

antibody and TCR molecules? What features of the amino acid
sequences in these regions are important for their functions?
• What are the differences in the types of antigens recognized
by antibodies and TCRs?
• What mechanisms contribute to the diversity of antibody and
TCR molecules? Which of these mechanisms contributes the
most to the diversity?
• What are some of the checkpoints during lymphocyte
maturation that ensure survival of the useful cells?
• What is the phenomenon of negative selection, and what is its
importance?
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