Immunoglobulin Structure and function.ppt

Immunoglobulin Structure and function

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Immunoglobulin
Structure & Function
Assigned Reading
Performance Ojectives
–Key terms
–Key Concepts
Content Outline
Short Answer Questions

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Angels & Devils

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Assigned Reading
Chapter: 5 pp107-142
Janis Kuby’s Immunology 3rd Ed

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Content Outline
Basic Structure
B Cell Receptor
Antigenic
Determinants of Igs
Immunoglobulin
Superfamily
Ig Classes
Monoclonal
Antibodies (MABS)
Immunoglobulin
Sequencing Studies
Immunoglobulin
Fine Structure

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Background

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Tiselius

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Electrophoresis

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General Structure of
Immunoglobulins
Electron microscopy
Crystallography
Hinge region
light & heavy chains
binding site
domains
fragments

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IgG Crystal

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X -Ray Crystallography
Crystallized Antigen
or Antigen -
Antibody Complex
Electron Density Map
of the Molecule
Three Dimensional
Images

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Antibodies contain a flexible
hinge region
EM evidence
Location
Chemical Structure
Advantages

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Antibody
Flexibility
EMs of same conc of
antibody mixed with
different conc of antigen
Ag-Ab complexes form
different shapes
Ag is inflexible
AB MUST BE FLEXIBLE

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Ag-Ab Complexes

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Immunoglobulin flexible
hinge

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Hinge Prolines

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Advantages of hinge
Rotation around central point
Bind antigens at various distances
Bind various ratios of antigens
Blocks or binds to antigens over a
wide range of concentrations

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Structure of IgG

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IgG: Domains

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IgG: Enzyme hydrolysis

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Ag-Ab complexes after Pepsin

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B Cell Receptor
BEFORE IMMUNIZATION
–Ig genes are organized & Igs assembled
–Anchored on membrane as specific
receptors for immunogen
AFTER IMMUNIZATION
–different expression of Ig genes
–lose anchor>> secreted
–different class of Ig secreted

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Antigenic Determinants of Igs
Immunoglobulins are immunogenic
–Inject human antibodies into a mouse
–Mouse develops anti-Human antibodies
–Use these antibodies to probe structure of
immunoglobulins
Three classes of anti-antibodies
–Isotypes
–Allotypes
–Idiotypes

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Antibody Classes: Outline
5 classes of antibodies
Concentration of classes
Structure and function of
–IgG
–IgM
–IgA
–IgD
–IgE

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Immunoglobulin Classes
Immunoglobulin A IgA,mA
Immunoglobulin D IgD, mD
Immunoglobulin E IgE, mE
Immunoglobulin G IgG,mG
Immunoglobulin M IgM,mM

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Immunoglobuin
concentrations

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Ig Superfamily

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IgG: Physicochemical
Properties
Molecular weight
–Sedimentation constant 7S
–Total 146,000
–Heavy chain 51,000
Isotypes 4
Heavy chain domains 4
Carbohydrate (%) 2-3
half life (days) 21

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Structure of IgG

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IgG: Domains

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IgG subclasses

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IgG: Functions
Bind antigen in tissue fluids
Promote destruction of antigen
–activates complement
Promote phagocytosis
–binds to Fc receptors on macrophages
Passive protection of fetus
–binds to receptors on placenta
–transferred to fetal circulation

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Functions of IgG Domains
V
H+ V
L Antigen binding
CH
1Complement binding c4b
CH
2complement binding C1q
CH
3Macrophage binding,Fc receptor
CH
2+CH
3neutrophil & K cell binding,
bind to placental syncytiotrophoblast

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Primary vs Secondary
Response

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IgM:Physicochemical
Properties
Molecular weight
–Sedimentation constant 19 S
–Total 900,000
–Heavy chain 65,000
Isotypes 1
Heavy chain domains 5
Carbohydrate (%) 12
half life (days) 5

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IgM: Structure

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IgM: Heavy chains

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IgM: Functions
Antigen receptor for B cell
Short term protection
–binds tightly to antigen
–(high avidity)

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IgA:Physicochemical Properties
Serum Secretory
Molecular weight
–Sedimentation constant7S 11S
–Total 160,000 385,000
–Heavy chain 52-56,000 52,56,000
Isotypes 2 2
Heavy chain domains4 4
Carbohydrate (%) 7-11 7-11
Half life (days) 6 6

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Structure of IgA

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IgA dimers

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EM of IgA

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Secretion of IgA

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IgA: Functions
Protects body surfaces against invading
microbes
Secretory IgA -
–transported to epithelium
–secreted in mucus, milk & tears
Blood IgA -
–transported to liver
–binds to hepatocytes
–secreted into bile

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J chain
Small polypeptide chain
found in polymeric classes of antibodies
Acts as a clasp to join prototype
subunits
produced by same cell that makes
antibodies

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J chain structure

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J chain: Beta sheets

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J Chain: Barrel

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IgE: Physicochemical
Properties
Molecular weight 188,000
–Sedimentation constant 8S
–Total 188,000
–Heavy chain 72,500
Isotypes 1
Heavy chain domains 5
Carbohydrate (%) 12
half life (days) 2

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Structure of IgE

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Molecular model of IgE

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IgE: Functions
Mediates inflammatory response
–Immunity to parasitic worms
–Mediates many allergies (type I)

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IgD Physicochemical
Properties
Molecular weight
–Sedimentation constant 7S
–Total 184,000
–Heavy chain 70,000
Isotypes 1
Heavy chain domains 4
Carbohydrate (%) 9-14
half life (days) 3

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IgD Structure

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IgD Hinge region

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Substructure of IgD

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Mouse IgD

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Mouse IgD

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IgD Function
Activation of B cells by antigen
–IgD : membrane antigen receptor
Indicator of Mature B cells
–low levels of IgD & IgM in Bone marrow
–high levels of IgD & IgM in peripheral
lymphoid organs

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DONE!!!

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Performance Objectives

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Key Terms
allotypes, allotypic determinants, Am
determinants, Gm determinants,
Km determinants, Bence Jones proteins,
Antibodies, immunoglobulin classes,
antibody subclasses, constant region
(C), Fab fragment, Fab' fragment,
F(ab')2 fragment, Fc fragment,
Framework residues (FR), Heavy chain,

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hinge regions, chimeric antibodies,
hybridoma, hypervariable regions (HVRs),
complematrarity determining
regions(CDRs)
Hypoxantinine-Aminoopterin-Thymidine
medium (HAT),idiotypes, idiotypic
determinants,
immunoglobulin domains
immunoglobulins, isotype, isotypic
determinants,

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joining chain (J), Light chain, monoclonal
antibodies, multiple myeloma,
Secretory Iga, Segmental flexibility,
variable regions (V)

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Key Concepts
Distinguish between the overall structure
and the fine structure of immunoglobulins
Describe the variable and constant regions of
immunoglobulins light and heavy chains
Explain the structural organization of the
variable regions of an immunoglobulins
light and heavy chains

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Differentiate between Hypervariable
regions and complementarity determining
regions
Contrast monoclonal antibody production
vs polyclonal antibody production
Discuss the differences in the biological
effector functions of the different classes of
immunoglobulins

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Diagram the procedure for monoclonal
antibody production
Construct a table comparing the
biological characteristics of the five
classes of immunoglobulins
Construct a table comparing the
biochemical and biophysical
characteristics of the five classes of
immunoglobulins

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Short Answer Questions

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The analysis of IgG molecules after
hydrolysis by pepsin and papain led to
similar yet different results. Explain.
What is the difference between and
immunoglobulin and a myeloma protein?
Why were myeloma proteins and Bence
Jones proteins critical to the early studies
on antibody structure? What has largely
replaced them?

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Differentiate among, complementarity
determining regions, hypervariable regions
and framework regions.
Explain the statements: "Antibodies can be
antigens" and "Antibodies can be used to
characterize antibodies".
Discuss the terms immunoglobulin
isotypes, immunoglobulin allotypes and
immunoglobulin idiotypes and give
examples of each.

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Compare conventional antibody production
with monocloanl antibody production and
recombinant antibody production.
What are antibody domains?
Why can't light chains be used to classify
antibodies?
The function of an immunoglobulin
molecule is related to its structure. Explain.

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Different cellular fusion products can
potentially occur in a hybridoma fusion.
–What are the potential products?
–Why are they a problem?
–How can you obtain the appropriate
combination?
Distinguish between idiotypes and anti-
idiotypes.