Download - B-Cell Epitopes
B-Cell Epitopes
Chapter 10
ClausLundegaard
Antibodies
Antibodies. What are they?
• Virtually any substance can elicit an antibody response. • Clear extra cellular pathogens• neutralizing antibodies
• Antibody repertoire• > 1011 in humans
• How is this possible?• ~ 30.000 genes in the humans genome!• Immunoglobulin gene rearrangement
Antibody Effect. Neutralizing Antibodies
Virus or Toxin
Neutralizing Antibodies Inhibit cellular infection
Clear pathogen infection
Antibody - Antigen interaction
Fab (fragment antigen binding)
Antigen
Epitope
Paratope
Antibody
B-Cells. How are they made?
B-lymphocytes each displaying a unique B-cell receptor
Stem Cell
Precurser B-lymphocytes
Gene rearrangements
Gene Shuffling
Number of gene segments
The 12/23 rule of recombinationrecombination signal sequence (RSS)
{
Only combined 12 RSS to 23 RSS
Mechanism of gene rearrangement
RAG proteins (recombination-activating genes)
Addition of P and N nucleotides
TdT: terminal deoxynucleotidyl transferase
Antibody variable regions, CDR’s (Complementarity-determining regions)
Variable regionsAlpha-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 chainCDR 1 = Light blueCDR 2 = CeriseCDR 3 = Yellow Light ChainCDR 1 = RedCDR 2 = GreenCDR 3 = Blue
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CDR Regions
CDR = complementarity determining region
http://212.219.234.139/html/anti_alpha.html
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.
Identification of CDR’s (II)
CDR-H1Start Approx residue 26 (always 4 after a Cys) [Chothia / AbM defintion];Kabat definition starts 5 residues laterResidues before always Cys-XXX-XXX-XXXResidues after always a Trp. Typically Trp-Val, but also, Trp-Ile, Trp-AlaLength 10 to 12 residues [AbM definition];Chothia definition excludes the last 4 residues
CDR-H2Start always 15 residues after the end of Kabat / AbM definition) of CDR-H1Residues before typically Leu-Glu-Trp-Ile-Gly, but a number of variationsResidues afterLys/Arg-Leu/Ile/Val/Phe/Thr/Ala-Thr/Ser/Ile/AlaLength Kabat definition 16 to 19 residues;AbM (and recent Chothia) definition ends 7 residues earlier
CDR-H3Start always 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-GlyLength 3 to 25(!) residues
Example
>BU02A02.1GVQCEVHLLESGGGLVQPGGSLRLSCAASGFTFYSYAMSWVRQAPGKGLEWVSANSGSGGSTYYADSVRGRFTISRDNSKNTLYLQMNSLSAEDTAVYFCAKAPGYYYYYGMDVWGQGTTVTVSSGKNGHSRAFV
15 amino acids after end of CDR1
Somatic hypermutations
B-Cell Activation (Proliferation depends on affinity) No
AffinityLow Affinity
No Affinity
High Affinity
Somatic Hypermutations
Memory B-cells
Plasma cells
B-Cell Activation
B Cell
T Helper Cell
Class II MHC
Bound Peptide
TCR
Cartoon by Eric Reits
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?
What did we find?
• Issue of next talk