Lecture 5: Antibody Structure

antibodies

produced when naïve B cells with surface Ig bind Ag causing activation to proliferate and differentiate into Ab secreting plasma cells

features of antibodies

• secreted Ig

• responds to extracellular pathogens and toxins

• binds pathogen to disable it or make it more susceptible to immune system

• circulates in blood plasma, lymph and stays at mucosal surfaces

structure of antibodies

• glycoprotein

• disulfide bonds

• hinge region: G, D, A

benefit of hinge region

makes more shapes possible to improve binding

classes/isotypes of Ab

• IgA

• IgD

• IgE

• IgG

• IgM

immunological domain

folded peptides that protect the antibody from degradation

hypervariable regions / complementarity determining regions

small regions of high amino acid sequence diversity within the V (variable) regions off Ig and TCR, which correspond with the antigen binding site on the light chain

benefit of hypervariable regions / complementarity determining regions

increases antigen binding diversity

affinity

strength of the antibody for its epitope

avidity

accumulated strength of multiple affinities summed by many binding interactions

multivalent antigen

antigen with >1 epitope that can be bound by Ab

transmembrane region

heavy chain

light chain

antigen binding site

variable region

linear epitope

binding sites are continuous on the epitope/protein amino acid sequence

discontinuous epitope

binding sites are separated on the epitope/protein amino acid sequence

benefit of B cell diversity

allows protection from a wide variety of pathogens

processes that contribute to B cell diversity

• VDJ/somatic hypermutation

• junctional diversity

• somatic hypermutation

processes that occur before antigen exposure

1. VDJ/somatic hypermutation

2. junctional diversity

3. alternative mRNA splicing (to co-express IgD and IgM)

4. allelic exclusion

processes that occur after antigen exposure

1. producing secreted antibodies

2. somatic hypermutation and isotype switching

3. effector function

recombination signal sequence (RSS)

a site for enzymes to identify to ensure correct directionality

structure of RSS

• 1 heptamer (7bp) + 12 bb spacer + nonamer (9bp)

• 1 heptamer (7bp) + 23 bb spacer + nonamer (9bp)

12/23 rule

only segments with opposite RSS will combine (12 + 23). this rule ensures that no D (diversity) regions are lost.

V(D)J recombinase

a set of enzymes that recombine V, D and J segments

structure of V(D)J recombinase

• recombination activating gene (RAG)

• terminal deoxynucleotidyl transferase

• DNA-PKcs and Ku70/80

• DNA ligase

• DNA Pol

recombination activating gene (RAG)

found only in lymphocytes, made up of 2 proteins (RAG 1 and RAG 2) that cuts DNA @ RSS

enzymes in V(D)J recombinase present in all nucleated cells that repair the DNA after it is cut

• terminal deoxynucleotidyl transferase

• DNA-PKcs and Ku70/80

• DNA ligase

• DNA Pol

production of IgM and IgD on the B cell surface with Igb and Iga

• BCR is membrane-bound and soluble antibody (IgM and IgD) is secreted

• Igb and Iga are required for surface-bound BCR to communicate inside the cell

allelic exclusion

allows only one heavy chain and one light chain per B cell

allelic exclusion benefits/functions

• prevents autoimmunity

• allows high avidity binding to more effectively clear pathogens

production of secreted IgM

• produced by alternative mRNA splicing with IgD

• it is often secreted first in humoral immunity

somatic recombination (SRC) / VDJ recombination / gene rearrangement

• a single gene segment of each type is brought together to form a DNA sequence encoding the V region of an immunoglobulin chain

• enzymes cut & rejoin DNA as directed by RSS

somatic recombination (SRC) / VDJ recombination / gene rearrangement benefit

increases diversity and specificity of antibody response

somatic recombination (SRC) / VDJ recombination / gene rearrangement steps

1. cut DNA: RAG complex binds two different RSS and cut

2. DNA repair: enzymes repair DNA forming signal joining and coding joint

somatic recombination (SRC) / VDJ recombination / gene rearrangement order of segment arrangement

D-J segments are joined first, then V joins D-J

coding joint

the DNA sequence formed by the joining of gene segments during V(D)J recombination

junctional diversification

random addition or subtraction of nucleotides at the coding joint

junctional diversification benefit

increases diversity and specificity of antibody response

isotype switching

by altering the heavy chain the isotype of the antibody is changed

isotype switching steps

1. AID replaces C with U

2. UNG removes U

3. APE-1 nicks both DNA strands

4. two switch regions are ligated together

5. the result is a new isotype

isotype switching benefit

improves specialization of function and ability to recruit effectors

somatic hypermutation

introduces point mutations by enzyme AID and then selects for antibodies that bind more tightly to the pathogen

somatic hypermutation benefit

improves Ag binding to Ab

affinity maturation

the increase of affinity of Ag binding sites due to SHM and selection of higher affinity BCRs

affinity maturation benefit

increases affinity of Ag binding sites

the only pentameric isotype

IgM

has (2 × 5=) 10 binding sites!

IgA function

neutralization

IgD function

works with IgM

IgE function

sensitization of mast cells (responsible for asthma + allergies)

IgG function

widest range of fucntion

IgM function

activates complement system

framework region

conserved, structurally stable portion of the variable domains of antibody heavy and light chains