Antibody Structure and Function - UTH between types of antibodies ... The V regions of every antibody differ from ... Iso, Allo, or Idio???
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Antibody Structure and Function
(Part I)
Keri C. Smith, Ph.D.January 14, 2014
Keri.C.Smith@uth.tmc.eduMSB 2.218
(or)
Anatomy and Physiology
of Antibodies
Overview
Physical properties of antibodies Structural and molecular features
Differences between types of antibodies Functional properties of antibodies
To review
Antibodies are gamma-globulins
Anatomy of Ig Light and Heavy Chains
Fab portion can bind antigenFc portion can be crystallized
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%
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
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
Domain Structure
Building blocks of all immunoglobulin superfamilydomains
Human Ig Domain Functions
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)
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
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)
The combination of H and L chain HV regions determines final antigen-specificity
Human Ig Domain Functions
Ig Hinge Regions
Short span of aabetween 1st and 2nd C domains
Rich in Cys and Pro Provides for flexibility
of the molecule
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
Fc region, multiple functions
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)
Features of Ig Isotypes
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
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
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
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.
Two Examples of Anti-Idiotypic Ab
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
Iso, Allo, or Idio???
Ig Structural Features
IgG Structural Features
IgG Structural Features II
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
IgG Subclasses (2)
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)
IgG Biological Properties (1)
Most common Ig in serum Distributed approximately equally between
intravascular and extravascular spaces Long half-life
IgG Recycling
Roopenian & Akilesh Nature Reviews Immunology 7, 715-725 (September 2007)
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
Three Roles for IgG
Antibody Dependent Cell-Mediated Cytotoxicity
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
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
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
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
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)
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