Structural and genetic diversity in antibody repertoires from diverse species

Abstract

The antibody repertoire is the fundamental unit that enables development of antigen specific adaptive immune responses against pathogens. Different species have developed diverse genetic and structural strategies to create their respective antibody repertoires. Here we review the shark, chicken, camel, and cow repertoires as unique examples of structural and genetic diversity. Given the enormous importance of antibodies in medicine and biological research, the novel properties of these antibody repertoires may enable discovery or engineering of antibodies from these non-human species against difficult or important epitopes.

Figure 1

V(D)J recombination and the structure of human and mouse antibodies. A schematic of the human heavy chain locus is shown, where multiple V_H, D_H, and J_H regions can rearrange in any given B-cell to produce a functional VDJ unit that encodes the heavy chain variable region. A close-up of the antigen binding region (bottom left) illustrates the regions encoded by the V, D, and J gene segments. A full length antibody is shown on the bottom right.

Figure 2

Evolutionary relationships among species: useful antibody innovations evolved several times in vertebrates. Deuterostome phylogeny highlights the divergence times between some lineages that are discussed in this review (in bold). Agnathans such as the lamprey have lymphocyte classes similar to jawed vertebrates, but employ diverse repertoires of variable lymphocyte receptors from somatically rearranged loci employing activation induced cytidine deaminase (AID). Gnathostomes such as the shark began using immunoglobulin superfamily receptors on lymphocytes and first evolved an immunoglobulin heavy chain in the absence of light chains. Birds use AID to gene convert an initially very restricted recombination activating gene (RAG) generated locus via sequence donation from variable segment pseudogenes. Camelids convergently evolved heavy-chain only antibody variants of IgG, and at least some members of the family Bovidae employ heavy chains with an ultralong CDR H3, creating another diverse disulfide stabilized domain. (Right) Structures of antibody binding fragments from diverse species. Ribbon diagrams of lamprey (PDB, 3E6J), shark (4HGK), chicken (4P48), camel (1ZVY), cow (4K3D), and human (1N8Z) antigen binding fragments. Key distinguishing features are that lampreys utilize a leucine rich repeat scaffold, as opposed to other species which use the immunoglobulin domain. Shark and camelid fragments are devoid of light chains, with camelids often using a longer disulfide-bonded CDR H3. Chickens have novel canonical classes of CDR H1, and disulfides in CDR H3. Cows have evolved an ultralong CDR H3 repertoire with a disulfide bonded knob that sits atop a β-ribbon stalk.

Figure 3

Novel processes for producing genetic diversity in chickens and cows. (a) Unlike humans or mice, chickens use a gene conversion strategy to create a diverse antibody repertoire. A single VDJ event occurs on the heavy chain locus, followed by gene conversion using sequence homology from a number of 5′ pseudogenes (ψV_H) which can be in rearranged form and donate diverse genetic fragments into the rearranged V region. (b) Ultralong CDR H3 and repertoire development in the cow. Cow antibodies with ultralong CDR H3s appear to use only one V_H region, V_HBUL, which recombines with a long D region, D_H2 to produce a germline VDJ recombined V-region. The single rearrangement then undergoes somatic hypermutation which may mutate residues to or from cysteine, changing the disulfide patterns of the repertoire, in addition to adding sequence diversity.

Figure 4

Protruding CDR H3 structures of camelid and cow antibodies. (a) CDR H3 structure and epitope binding of camel antibody fragments. (Left) Closeup of the structure of a camelid Fv, showing the long CDR H3 loop with a disulfide bond between CDR H3 and CDR H1. Unique epitopes bound by camelid Fvs, including the β-adrenergic GPCR (middle) and enzymatic active site of lysozyme (right). Note the protruding CDR H3 which reach into concave epitopes on each of these antigens. (b) Structures of two cow Fab fragments, BLV1H12 and BLV5B8, showing ultralong CDR H3s that are composed of a β-ribbon ‘stalk’ and disulfide bonded ‘knob’ structural minidomains. (c) Closeup views of the surface of the knob domains of BLV1H12 and BLV5B8. The sequences of the knobs are shown below, and a two-dimensional representation of the loops are shown to the right. The disulfide patterns of the two knobs are different, as indicated above the sequences and in the two dimensional loop schematic.