Dr. Nabil MTIRAOUI, M.Sc, Ph.D

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Immunoglobulin
Structure and Function

• Dr. Nabil MTIRAOUI, M.Sc, Ph.D

Lecture 8
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Definition and Properties
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Outline of Lecture

1. Recognize the structure of immunoglobulin
   molecule
2. Know the different types of immunoglobulin.
3. Understand the biological activities of each
   type.
4. Differentiate between types of immunoglobulin.
5. Define isotype switching and antibody diversity.
6. Differentiate between types of FC receptors.

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Antibodies (or Immunoglubulins)

• The chemical information of immunoglobulin was
  provided by Tiselius and Kabat in the early
  1940s.
• In 1950s, Porter and Edelman revealed the basic
  structure of immunoglobulin molecule.
• Antibodies are products of antigen- activated B-
  lymphocytes.
• They are the main effectors of humoral immunity.

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Definition

• An antibody or immunoglobulin (Ig) is a
  glycoprotein that is made by plasma cells in
  response to an antigen and can recognize and bind
  to the antigen that caused its production.
• Antibodies are produced by B cell

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General Functions of Immunoglobulins

• Ag binding
• Can result in protection
• Valency
• Effector functions
• Fixation of complement
• Binding to various cells
• Usually requires Ag binding

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Basic Immunoglobulin Structure

• Fab region
• Fc region
• Heavy chain with one variable (VH) domain
• followed by a constant domain (CH1), a hinge
  region, and two more constant (CH2 and CH3)
  domains.
• Light chain with one variable (VL) and one
  constant (CL) domain
• Antigen binding site (paratope) It is the area
  of Ig molecules that interacts specifically with
  epitope of the Ag
• Hinge regions.

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Basic Immunoglobulin Structure

• Abs have more than one antigen combining site
  Some bivalent Ab molecules can combine to form
  multimeric Abs that have up to 10 combining
  sites.
• All immunoglobulin have a basic structure
  composed of 4 polypeptide chains connected to
  each other by disulfide bonds.
• Each light chain consist of 220 Aa and has a mass
  of approx. 25kDa.
• Each heavy chain consists of about 440 Aa and has
  a mass of 50-70kDa.

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Basic Immunoglobulin Structure

• Both light and heavy chains contain 2 different
  regions
• constant and variable region
• The four chains are arranged in the form of a
  flexible Y with the hinge region and is termed
  as crystallizable fragment (Fc) and contains the
  site at which Ab binds.
• Top of the Y consist of two Ag binding
  fragments (Fab) that bind with antigenic
  determinant sites.
• The four chains are linked by disulfide bonds.

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Basic Immunoglobulin Structure

• Light chain
• The light chain may be either of two distinct
  forms called Kappa and Lambda and can be
  distinguished by aa sequence of carboxyl portion
  of the chain.
• Heavy chain
• In the heavy chain NH2 terminal has a pattern of
  variability similar to that of kappa and lambda
  of the light chain.

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The variable (V) regions.

• The first 100 or so amino acids at the N-terminal
  of both H and L chains vary greatly from antibody
  to antibody.
• These are the variable (V) regions.
• The amino acid sequence variability in the V
  regions is especially pronounced in 3
  hypervariable regions.
• Together they construct the antigen binding site
  against which the epitope fits
• Only a few different amino acid sequences are
  found in the C-terminals of H and L chains.
• These are the constant (C) regions.

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The variable (V) regions.
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The Hypervariable regions.

• Hypervariable regions In the variable regions of
  both LH chains, there are 3 extremely
  hypervariable amino acids sequences that form the
  Ag binding sites.
• The hypervariable regions form the region
  complementary in structure to the epitope. These
  regions are involved in the formation of paratope.

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The constant (C) regions.

• two different kinds of C regions for their L
  chains producing
• kappa (?) L chains
• lambda (?) L chains
• five different kinds of C regions for their H
  chains producing
• mu (µ) chains (the H chain of IgM antibodies)
• gamma (?) chains (IgG)
• alpha (a) chains (IgA)
• delta (d) chains (IgD)
• epsilon (e) chains (IgE)

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Fc and Fab regions

• The proteolytic enzyme papain breaks each Ig
  molecule into 3 fragments at the hinge region.
• The single crystallizable fragment (Fc region)
  includes part of the constant domain that
  occupies the stem.
• There are 2 antigen-binding fragments (Fab
  region), which include the entire light chain and
  variable and constant portions of the heavy
  chain.
• Ig G

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Fc and Fab regions

• Antigen-binding fragment (Fab)
• recognize Ag
• contain Ig idiotype
• Unique protein sequence that identifies each Ab
• Crystallizable fragment (Fc)
• define isotype of Ig
• bind FcR for all functional attributes of Ab
• Fab link to Fc by hinge region

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Fonction of Fc and Fab regions

• By binding to specific proteins the Fc region
  ensures that each antibody generates an
  appropriate immune response for a given antigen.
• The Fc region also binds to various cell
  receptors, such as Fc receptors, and other immune
  molecules, such as complement proteins.
• Thus, Ab mediates different physiological effects
  including opsonization, cell lysis, and
  degranulation of mast cells, basophils and
  eosinophils.

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Activation of complement

• Antibodies that bind to surface antigens on, for
  example a bacterium, attract the first component
  of the complement cascade with their Fc region
  and initiate activation of the "classical"
  complement system
• This results in the killing of bacteria in two
  ways
• First, the binding of the antibody and complement
  molecules marks the microbe for ingestion by
  phagocytes in a process called opsonization
• Secondly, some complement system components form
  a membrane attack complex to assist antibodies to
  kill the bacterium directly.

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Types of FC receptors
FC receptor Affinity to Ig Cell distibution Function
Fc?Rl (CD64) High binds IgG1a and IgG3 MQ. Neutrophils and eosinophils Phagocytosis, activation of phagocytes
Fc?RllA (CD32) Low MQ. Neutrophils and eosinophils,platelet Phagocytosis,cell activation (inefficient)
Fc?RllB (CD32) Low B lymphocytes Feed back inhibition of B cells
Fc?RlllA (CD16) Low NK ADCC
FceRl High bind monomeric IgE Mast cells, basophils, eosinophils Avtivation, degranulation of mast cell, basophils
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CLASSES (ISOTYPES) OF IMMUNOGLOBULINS

• Classes based on constant region of heavy chains
• Immunoglobulin A (IgA)
• Immunoglobulin D (IgD)
• Immunoglobulin E (IgE)
• Immunoglobulin G (IgG)
• Immunoglobulin M (IgM)
• Differentiation of heavy chains
• Length of C region, location of disulfide bonds,
  hinge region, distribution of carbohydrate
• Classes have different effector functions

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CLASSES (ISOTYPES) OF IMMUNOGLOBULINS
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Immunoglobulin G

• Structure
• Monomer (7S)

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IgG

• It is the major Ig in normal serum, accounting
  for 75 of the total Ig pool. It is a monomeric
  unit (2 heavy chain 2 light chain), MW 160,000.
  It can bind 2 Ag molecules. 4 subclasses are
  known IgG1,2,3,4. Its biological activities
  include
• Its half life time is 23 days and is the longest
  of all Igs.
• It is the major Ab in the secondary immune
  response.
• It is the only Ab that can cross placenta (IgG2
  does not cross well) and provide immunity to the
  newborn during the first months after birth.
  Transfer is mediated by a receptor on placental
  cells for FC region of IgG .
• It diffuse into the extravascular neutralizing
  bacterial toxins (antitoxin).
• It enhance phagocytosis (opsonization) by coating
  bacteria and attaching by its FC portion to FC
  receptor on phagocytic cells.
• It can fix and activate complement (by IgG1 and
  IgG3)

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Immunoglobulin A
Secretary Ab First line defense for microbes

• IgA a doublet guards the entrance to the body.
  170,000 MW in serum and 400,000 MW in external
  secretions, 15 of Ig in serum, found in the
  blood as a monomer, and in tears, saliva,
  colustrum, nasal, vaginal, prostatic and
  bronchial secretions as a dimer.
• Blocks attachment of microbes to mucous
  membranes
• It concentrates in body fluids such as tears,
  saliva, and secretions of the respiratory and
  gastrointestinal tracts.

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IgA

• Structure
• Serum - monomer
• Secretions (sIgA)
• Dimer (11S), sIgA molecule consists of two H2L2
  units plus one molecule each of J chain and
  secretory component (SC or SP)

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IgA

• Properties
• 2nd highest serum Ig
• Major secretory Ig (saliva, tears, respiratory,
  intestinal, and genital tract secretions.)
• Does not fix complement unless aggregated
• Binds to Fc receptors on some cells

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Immunoglobulin M
Macroglobulin primary immune response Bacteriolyti
c

• IgM usually combines in star-shaped clusters.
  pentamer,
• It tends to remain in the bloodstream, 10 of
  blood Ig,
• found on the surface of B lymphocytes.
• Activates the complement system.

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IgM

• Structure
• Pentamer (19S)
• composed 5 H2L2 units plus one
  molecule of
• J chain
• Extra domain (CH4)
• J chain

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Fixation of C1 by IgG and IgM Abs
No activation
Activation
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IgM

• Structure
• Properties
• 3rd highest serum Ig
• First Ig made by fetus and B cells
• Produced early in the primary response
• The most efficient Ig
• Fixes complement

Agglutinating Ig Binds to Fc receptors B cell
surface Ig
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Immunoglobulin D
Membrane bound antibody Found on B-cell
membrane Memory function

• 180,000 MW, activity is not well known. 0.2 of
  plasma Ig. 13 carbohydrate content. Also found
  on the surface of B lymphocytes, where it somehow
  regulates the cell's activation

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IgD

• Structure
• Monomer
• Tail piece

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IgD

• Structure
• Properties
• 4th highest serum Ig
• Expressed in B cell surface Ig
• Does not bind complement
• FUNCTIONS
• B cell activation .
• Acts as receptor for Ag binding.

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Immunoglobulin E
Activate mast cells Release vasoactive
amines Respond to allergens Mediate
hypersensitivity reactions

• 90 000 MW, largest immunoglobulin, present in
  extremely low levels in a healthy individual.
• IgE levels rise in response to parasitic
   infections and in  allergic reactions.
• Bind and activate mast cells. Mast cells cause
  acute inflammatory response (e.g. swelling,
  redness, pain and itchiness). 
• Hay fever is a condition caused by too much IgE
  activity.

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IgE

• Structure
• Monomer
• Extra domain (CH4)

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IgE

• Structure
• Properties
• Least common serum Ig
• Allergic reactions
• Parasitic infections
• Does not fix complement
• FUNCTIONS
• Responsible for immediate hypersensitivity or
  allergic reactions.
• Binds to Fc receptors on basophils and mast
  cells.
• Release of substances like histamine , bradykinin
  and other vasoactive mediators.

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Properties of immunoglobulins
IgG IgA IgM IgD IgE
1. Serum conc. () 85 5-15 5-10 lt1 lt1
2. Mol. Wt. 160,000 170,000 385,000 960,000 184,000 188,105
3.Sed. coeff. 7S 7S 19S 7S 8S
4.Heavy chain class Gamma Alpha Mu Delta Epsilon
5.Light chain K L K L K L K L K L
6. Valency 2 2 or multiple of 2 5 (10) 2 2
7.No of basic 4-polypeptide chains Monomer Dimer or Trimer Pentamer Monomer Monomer
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IgG IgA IgM IgD IgE
8.Placental transport _ _ _ _
9.Present in milk _ _ _
10.Selectie secretion by seromucus gland _ _ _ _
11. Intravascular distribution() 45 42 80 75 50
12.Carbohydrate () 3 11 10 13 12
8.Subclasses IgG1-4 IgA1-2 _ _ _
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IgG IgM IgA IgD IgE
14.Comple-ment fixation
A.Classical _ _ _
B.Alternati ve _ _ _ _
15.Half life (days) 23 6 5 2-3 2-3
16.Princip-al site of action Serum Secretion Serum Receptor for B cells Mast cells
17.charact-eristic propert-ies precipitins, antitoxins, compleme-nt fixation, late Ab Serum and secretory Abs Agglutinin, opsonin , lysin , early Ab Not known (B-cell activation) Reaginic Ab (anaph- ylaxis)
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Thank You