antibodies Flashcards
production of antibodies
humoral and cellular components
b cells are lymphocytes central to our humoral response
b cells originate and mature in bone marrow
2 types of b cells
antibody secretory cells or memory b cells
what do plasma cells secrete
antibodies of the same specificity as the membrane bound immunoglobulin expressed by their b cell precursor
what do stimulated b cells give rise to
antibody secreting cells - bacterium have unique binding sites that bind with b cell receptors
draw the structure of an Ig
check notes lol
Fab regions of Igs
fragement antigen binding - responsible for antigen binding
Fc regions if Igs
fragment crystallisable - responsible for effector function
what does the flexible hinge of IgG allow
allows both arms to bind many arrangements of antigens on pathogen surfaces
hinge is key to the structure and gives it its function, give antibodies flexible arms to move idependently of one another
allows binding of both arms at once and increases strength of binding
types of light chains
lambda and kappa - both light chains are the same on individual Igs
what determines the class of the antibody
the heavy chain
what are the globular domains of antibodies comprised of
2 beta sheets with a total of 7 or 8 antiparallel b strands linked by a disulphide bridge to form a beta barrel, antiparallel sheets are connected by loops
how do different antibodies exists
variability in loop domains of VH and VL
what part of the antibody forms the antigen binding site
hypervariable loops commonly termed complimentary determining regions (CDRs)
form close lying loops at the end of the v domain
when VH and VL domains are paired their 6 cdrs create an antigen binding site
antigen binding sites have complimentary 3d structures to the antigenic epitope providing high specificity and high affinity
how is antibody diversity achieved?
any substance can elicit an antibody response
responses to simple substances are diverse
Ig genes are organised different to normal genes allowing vast antibody responses
in all cells bar b cells, Ig genes are in fragmented form that cannot be expressed
Ig heavy and light chain loci consist of families of gene segments arrayed sequentially along the chromosome
inherited in this form through egg and sperm - germline configuration
in developing b cells, Ig gene segments must be rearranged to assemble functional light and heavy chains
multiple variable region gene segments
VH domain is encoded by V, D, J gene segments
VL domain is encoded by V, J gene segments
j = joining
d = diversity
when does Ig gene rearrangement occur
heavy chains = starts in early pro B cells
light chains + in small pre B cell
how do pro b cells randomly recombine heavy chain gene segments
dna is spliced out during early development of b cells
1. d gene segment is joined to a J gene segment
2. v gene segment is joined to a DJ gene segment
events are called somatic recombinants because they occur in cells other than germ cells
section of intervening DNA needs to be cut out and los tto allow juxtaposition
splice and loss is irreversible
kappa light chain rearrangement
repeated light chain rearrangement is possiblw
guided by conserved non coding sequences known as recombination signal sequences
12.23 rule
recombination signal sequences flank recombination sites
recombination signal sequences consist of a conserved heptamer, non conserved spacer (12 or 23 bp) and conserved monomer
genes with 12bp spacer can recombine with 23bp spacer
enzymatic requirements for recombination
V(D)J recombinase - recombine V,D and J genes
RAG1 and RAG2 made in lymphocytes, they associate with eachother and other proteins known as high mobility group of proteins to give the RAG complex
other components that are able to: repair double stranded DNA, bend DNA, modify ends of broken DNA
these include: DNA ligase 4, DNA dependent protien kinase, arternis and KV protein
process of antibody gene recombination
one rag complex bins 23 spacer RSS another binds 12 spacer RSS
rag complex interactions align RSS and cleaves DNA at ends of the gene segments
DNA hairpin is formed at each end of the gene segment and a clean break occurs at the heptamer ends
rag complexes hold dna in place while broken ends are rejoined by dna repair enzymes - non homologous end joining
coding joint formed in chromosome, signal joint in removed piece of circular DNA
generation of junctional diversity
enzymes that open the coding joint introduce additional sequence diversity into CDR3 in several ways
ways to generate junctional diversity
rag complex cleaves the heptamer RSSs from D and J gene segments to yield DNA hair pins - the nick that opens the hairpin can occur at several diff positions
RAG complex opens the hairpins by nicking one strand of DNA generating palindromic P nucleotides - forms a single stranded end which complements the bases that were originally on 2 dna strands now on the same strand, this creates a sequence that would form a palindrome in dsDNA
pairing of strands - once sequence complementarity allows strands to pair, single stranded gaps are filled in with complimentary nucleotides
unpaired nucleotides are removed by an exonuclease - contribution of P and N nucleotides to resulting amino acids sequence in CDR3 is called junctional diversity
gaps are filled in by dna synthesis and ligation to form a coding joint
diversity of Ig repertoire is gereated by 4 main processes
- combinational diversity - many copies of each gene segment typw combine in different ways
- junctional diversity - nucleotides added or subtracted at joins between gene segments during enzymatic steps
- many combinations of light and heavy chains, v regions pair to form antigen binding sites
- somatic hypermtutation - introduces point mutations into rearranged v region genes of activated b cells
human heavy chain Ig locus
Ch genes form clusters 3’ to Jh
each Ch has multiple exons each encoding an individual Id domain
Cmieu closest to Jh gene segments and first complete transcript is produced
subsequently additional isotypes are generated by irreversible change in DNA processes known as isotype switching
how do developing and naive b cells express IgM and IgD
they use alternative splicing,
circulating b cells that havent encountered antigens are naive b cells - they express IgM and IgD on the surface
this is accomplished by splicing of the primary RNA transcript
how do b cells produce Ig of single antigen specificity
Ig gene rearrangement is tightly controlled
ensures only 1 light and 1 heavy chain is expressed
this is called allelic exclusion
what is the first class of Ig that is secreted
IgM
what are Ig first form
membrane bound form present on B cell surfaces
what are b cell receptors made up of
cell surface IgM with invariant Ig alpha and Ig beta proteins
what are Ig invariant chains important for
signalling once antigens bind to IgM
how do Igs anchor to the antigen membrane
heavy chain hydrophobic sequences near terminus anchor them into the membrane
transmembrane Ig have hydrophobic transmembrane domain of 25 amino acids that act as the anchor
how are membrane and secreted forms of Ig generated
by alternate processing of the heavy chain
secretory Ig has a secretory tail, transmembrane Ig have hydrophobic tails for anchors
2 carboxy termini are encoded in separate exons and are selected by alternative RNA processing
