antibody structure and function - uth between types of antibodies ... the v regions of every...
TRANSCRIPT
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Antibody Structure and Function
(Part I)
Keri C. Smith, Ph.D.January 14, 2014
[email protected] 2.218
(or)
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Anatomy and Physiology
of Antibodies
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Overview
Physical properties of antibodies Structural and molecular features
Differences between types of antibodies Functional properties of antibodies
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To review
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Antibodies are gamma-globulins
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Anatomy of Ig Light and Heavy Chains
Fab portion can bind antigenFc portion can be crystallized
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Ig Light Chains
Two types of L chains are found in Ig of all animals, aa sequence differs
Kappa chains-Human 60%, mice 95% Lambda chains-Human 40%, mice 5%
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Ig Heavy Chains
5 classes of H chains in humans Similarities in aa sequence, but each class has a
unique sequence H chains named with Greek letters
corresponding with the class name, IgG, IgA, IgM, IgE, IgD (γ, α, μ, ε, δ)
IgG has 4 subclasses, IgA has 2 subclasses
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Specific Domains All classes of Ig Each 100-110 aa has a 60 aa S-S loop V domains code the paratope, binds Ag C domains code regions important for mediating
secondary biological functions
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Domain Structure
Building blocks of all immunoglobulin superfamilydomains
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Human Ig Domain Functions
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Ig Variable and Hypervariable Regions
The V regions of every antibody differ from those of every other Ab
Sequence variability is concentrated in certain segments of both L and H chain V regions
3 Hypervariable (HV) regions in both L and H chain V regions.
These are epitope contact aa regions, CDR (complementarity determining regions)
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The ratio of the number of different amino acids at a given position to the frequency of the most common amino acid at that position is defined as VARIABILITY
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Formation of the CDR
Regions between HV regions are framework regions (FR)
FR form β sheets, HV regions on outer edges of barrel
Total of 6 HV loops = complementarity-determining region (CDR)
(So-called because the surface that is formed is complementary to the antigen that is bound)
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The combination of H and L chain HV regions determines final antigen-specificity
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Human Ig Domain Functions
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Ig Hinge Regions
Short span of aabetween 1st and 2nd C domains
Rich in Cys and Pro Provides for flexibility
of the molecule
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Fc portion
The portion of the Ab that allows for interaction of immune complexes with other cells
Many different forms of Fc receptors (FcR) exist May be regulatory or stimulatory
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Fc region, multiple functions
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Ig Classes or Isotypes
Result from structural variation in the heavy chains
Are 5 major classes or isotypes of Ig in Humans IgG, IgA, IgM, IgE, IgD Subclasses: IgG (4) and IgA (2)
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Features of Ig Isotypes
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Ig Allotypes
Allelic variants of Ig Constant regions Co-dominant autosomal Mendelian genes Are allotypic variants of γ, α, and κ chains Allotypes used forensically Allotype may confer some biological
advantage for some infectious agents
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Human Kappa Light Chain Allotypes
ALLOTYPE AMINO ACID DIFFERENCES
Km(1) Val @ 153, Leu @ 191
Km(1,2) Ala @ 153, Leu @ 191
Km(3) Ala @ 153, Val @ 191
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Allotype effects in infection and autoimmunity
Children with G2m(23) allotype had higher levels of immunity to H. influenzae type B than G2m(23)-negative children and the Km(1) allotype was found more effective than the other kappa chain allotypes. (Ambrosino, et al. 1985, J. Clin. Invest. 75:1935; Granoff, et al. 1986, J. Inf. Dis. 154:257.)
Immunoglobulin GM and KM genotypes in Korean patients with systemic lupus erythematosus. Lee et al. Rheumatol Int. 2007 Jan;27(3):219-24
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Ig Idiotypes
Idios: Greek, unique for Ab of one specificity. Are epitopes in Ig V regions Provide a natural autoimmune mechanism for
regulating Ab levels Anti-id antibodies are routinely found and down-
regulate production of Ab.
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Two Examples of Anti-Idiotypic Ab
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Clinical Relevance of Idiotypes
Active immunotherapy with 1E10 anti-idiotype vaccine in patients with small cell lung cancer: report of a phase I trial.
Neninger et al. Cancer Biol. Ther. 2007. 6:145
Anti tumor mAb
Anti- Anti tumor mAb
Anti-Anti –anti-tumor mAb
Also reacts with tumor antigen
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Iso, Allo, or Idio???
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Ig Structural Features
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IgG Structural Features
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IgG Structural Features II
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IgG Subclasses
IgG1, IgG2, IgG3, IgG4 Have about 90% homology in their amino acid
sequences Differ in the numbers and arrangement of the
interchain disulfide bonds This results in differences in their functional
properties
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IgG Subclasses (2)
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Important Differences Between Human IgG Subclasses
IgG1 IgG2 IgG3 IgG4
Occurrence (% of total IgG) 70 20 7 3
Half-life 23 23 7 23
Complement binding + + +++ —
Placental passage ++ ± ++ ++
Binding of monocytes +++ + +++ ±
IgG Subclasses (3)
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IgG Biological Properties (1)
Most common Ig in serum Distributed approximately equally between
intravascular and extravascular spaces Long half-life
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IgG Recycling
Roopenian & Akilesh Nature Reviews Immunology 7, 715-725 (September 2007)
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IgG Biological Properties (2)
Agglutination and Precipitation Placental passage (G1, G3, G4) Opsonization Antibody dependent cell mediated cytotoxicity
(G1, G3) Complement Activation (G1, G2, G3+) Toxin/Viral Neutralization--Passive Ab infusion Bacterial immobilization
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Three Roles for IgG
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Antibody Dependent Cell-Mediated Cytotoxicity
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Clinical Correlation
Hemolytic disease of the newborn. Case #45 Mom is Rh-. Dad is Rh+. 2 normal pregnancies. 3rd pregnancy:
increased indirect Coombs titer (reaction of mom’s Ab with Rh+ red cells)
Increased billirubin in amniotic fluid Low hematocrit in umbilical cord blood
Treatment = Infusion of fetus with type O Rh-RBC
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Mechanism of fetal RBC destruction
Maternal IgG antibodies specific for Rh are actively transported acrossthe placenta
Opsonize fetal Rh+ RBC for phagocytosis by liver cells
Fetal hematocrit drops to dangerous or fatal levels. Prophylactic treatment = Rhogam
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IvIg
Purified, polyclonal Igpooled from plasma of thousands of donors
In use since the 1980s, originally for immunodeficiency disorders
Also being used in: Solid organ transplantation,
Wegener's granulomatosis,vasculitis, heart failture, spinal chord injury, etc……..
NEJM 2001, Volume 345:747-755
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How does IvIg work?
Inhibition of complement activation?
Direct effects on B cells T cells (Treg) Dendritic cells Macrophages
FcRIIb binding?
Trends in Immunology Volume 29, Issue 12, December 2008, Pages 608-615
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IgG Review
Gamma Globulin, Enzymatic Fragmentation
L & H Chains, V & C Domains, Hinge Regions, Variability and Hypervariable regions
4 Isotypes (Classes)
Allotypes, Idiotypes
IgG Anatomy: γ H Chains, κ/λ L Chains
IgG Physiology: Agglutination, Placental Passage, Opsonization, ADCC, Complement Binding, Toxin/Viral Neutralization, Bacterial Immobilization, Recycling (long half life)