B-Cell Epitopes

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B-Cell Epitopes

• Chapter 10

Claus Lundegaard
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Antibodies
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Antibodies. What are they?

• Virtually any substance can elicit an antibody
  response.
• Clear extra cellular pathogens
• neutralizing antibodies
• Antibody repertoire
• gt 1011 in humans
• How is this possible?
• 30.000 genes in the humans genome!
• Immunoglobulin gene rearrangement

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Antibody Effect. Neutralizing Antibodies
Virus or Toxin
Neutralizing Antibodies
Inhibit cellular infection Clear pathogen
infection
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Antibody - Antigen interaction
Antigen
Fab (fragment antigen binding)
Epitope
Antibody
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B-Cells. How are they made?
Stem Cell
Precurser B-lymphocytes
Gene rearrangements
B-lymphocytes each displaying a unique B-cell
receptor
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Gene Shuffling
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Number of gene segments
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The 12/23 rule of recombination
recombination signal sequence (RSS)

Only combined 12 RSS to 23 RSS
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Mechanism of gene rearrangement
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RAG proteins (recombination-activating genes)
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Addition of P and N nucleotides
TdT terminal deoxynucleotidyl transferase
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(No Transcript)
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Antibody variable regions, CDRs
(Complementarity-determining regions)
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CDR Regions
Variable regions Alpha-carbon trace of the
structure of the heavy chain and light chain
variable regions of a typical antibody. The
framework regions of both chains are shown in
grey whilst the complementarity determining
regions (CDRs) are coloured individually,
i.e. Heavy chain CDR 1 Light blue CDR 2
Cerise CDR 3 Yellow Light Chain CDR 1
Red CDR 2 Green CDR 3 Blue
CDR complementarity determining region
http//212.219.234.139/html/anti_alpha.html
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Identifying CDR regions

• The Kabat definition is based on sequence
  variability and is the most commonly used
• The Chothia definition is based on the location
  of the structural loop regions
• The AbM definition is a compromise between the
  two used by Oxford Molecular's AbM antibody
  modelling software
• The contact definition has been recently
  introduced by us and is based on an analysis of
  the available complex crystal structures.

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Identification of CDRs (II)
CDR-H1 Start Approx residue 26 (always 4 after a
Cys) Chothia / AbM defintion Kabat definition
starts 5 residues later Residues before always
Cys-XXX-XXX-XXX Residues after always a Trp.
Typically Trp-Val, but also, Trp-Ile,
Trp-Ala Length 10 to 12 residues AbM
definition Chothia definition excludes the last
4 residues CDR-H2 Start always 15 residues after
the end of Kabat / AbM definition) of
CDR-H1 Residues before typically
Leu-Glu-Trp-Ile-Gly, but a number of
variations Residues after Lys/Arg-Leu/Ile/Val/Phe/
Thr/Ala-Thr/Ser/Ile/Ala Length Kabat definition
16 to 19 residues AbM (and recent Chothia)
definition ends 7 residues earlier CDR-H3 Start a
lways 33 residues after end of CDR-H2 (always 2
after a Cys) Residues before always Cys-XXX-XXX
(typically Cys-Ala-Arg) Residues after always
Trp-Gly-XXX-Gly Length 3 to 25(!) residues
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Example

• gtBU02A02.1
• GVQCEVHLLESGGGLVQPGGSLRLSCAASGFTFYSYAMSWVRQAPGKGLE
  WVSANSGSGGSTYYADSVRGRFTISRDNSKNTLYLQMNSLSAEDTAVYFC
  AKAPGYYYYYGMDVWGQGTTVTVSSGKNGHSRAFV

15 amino acids after end of CDR1
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Somatic hypermutations
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B-Cell Activation (Proliferation depends on
affinity)
No Affinity
Low Affinity
No Affinity
Somatic Hypermutations
High Affinity
Plasma cells
Memory B-cells
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B-Cell Activation
T Helper Cell
TCR
B Cell
Class II MHC
Bound Peptide
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Cartoon by Eric Reits
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Controversial issues

• Is the 11/23 rule always obeyed?
• Can you have multiple D genes?
• Can D genes be inserted backwards?
• I.e can both D and the inverted D genes be used?
• Does V, D and J palindrom segments exits?

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What did we find?

• Issue of next talk