The Unusual Genetics and Biochemistry of Bovine Immunoglobulins.

Antibodies are the key circulating molecules that have evolved to fight infection by the adaptive immune system of vertebrates. Typical antibodies of most species contain six complementarity-determining regions (CDRs), where the third CDR of the heavy chain (CDR H3) has the greatest diversity and often makes the most significant contact with antigen. Generally, the process of V(D)J recombination produces a vast repertoire of antibodies; multiple V, D, and J gene segments recombine with additional junctional diversity at the V-D and D-J joints, and additional combinatorial possibilities occur through heavy- and light-chain pairing. Despite these processes, the overall structure of the resulting antibody is largely conserved, and binding to antigen occurs predominantly through the CDR loops of the immunoglobulin V domains. Bovines have deviated from this general paradigm by having few VH regions and thus little germline combinatorial diversity, but their antibodies contain long CDR H3 regions, with substantial diversity generated through somatic hypermutation. A subset of the repertoire comprises antibodies with ultralong CDR H3s, which can reach over 70 amino acids in length. Structurally, these unusual antibodies form a β-ribbon "stalk" and disulfide-bonded "knob" that protrude far from the antibody surface. These long CDR H3s allow cows to mount a particularly robust immune response when immunized with viral antigens, particularly to broadly neutralizing epitopes on a stabilized HIV gp140 trimer, which has been a challenge for other species. The unusual genetics and structural biology of cows provide for a unique paradigm for creation of immune diversity and could enable generation of antibodies against especially challenging targets and epitopes.

Stanfield RL ，Haakenson J ，Deiss TC ，Criscitiello MF ，Wilson IA ，Smider VV ... -  《-》

Diversity in the Cow Ultralong CDR H3 Antibody Repertoire.

Haakenson JK ，Huang R ，Smider VV 《Frontiers in Immunology》

A Broad Role for Cysteines in Bovine Antibody Diversity.

Ab diversity in most vertebrates results from the assortment of amino acid side chains on CDR loops formed through V(D)J recombination. Cows (Bos taurus) have a low combinatorial diversity potential because of a small number of highly homologous V, D, and J gene segments. Despite this, a subset of the Ab repertoire (∼10%) contains exceptionally long CDR H chain (HC) 3 (H3) regions with a rich diversity of cysteines and disulfide-bonded loops that diversify through a single V-D-J recombination event followed by massive somatic hypermutation. However, the much larger portion of the repertoire, encoding shorter CDR H3s, has not been examined in detail. Analysis of germline gene segments reveals noncanonical cysteines in the HC V regions and significant cysteine content in the HC D regions. Deep sequencing analysis of naturally occurring shorter CDR H3 (<40 aa) Ab genes shows that HC V and HC D regions preferentially combine to form a functional gene with an even number of total cysteines in the final V region, suggesting that disulfide bonds contribute to diversity not only in ultralong CDR H3 bovine Abs but in shorter CDR H3 bovine Abs as well. In addition to germline "hard-coded" cysteines, the bovine Ab repertoire can produce additional cysteine codons through somatic hypermutation, further diversifying the repertoire. Given the limited combinatorial diversity at the bovine Ig loci, this helps to explain how diversity is created in shorter CDR H3 Abs and potentially provides novel structural paratopes in bovine Ab combining sites.

Haakenson JK ，Deiss TC ，Warner GF ，Mwangi W ，Criscitiello MF ，Smider VV ... -  《-》

Reshaping antibody diversity.

Some species mount a robust antibody response despite having limited genome-encoded combinatorial diversity potential. Cows are unusual in having exceptionally long CDR H3 loops and few V regions, but the mechanism for creating diversity is not understood. Deep sequencing reveals that ultralong CDR H3s contain a remarkable complexity of cysteines, suggesting that disulfide-bonded minidomains may arise during repertoire development. Indeed, crystal structures of two cow antibodies reveal that these CDR H3s form a very unusual architecture composed of a β strand "stalk" that supports a structurally diverse, disulfide-bonded "knob" domain. Diversity arises from somatic hypermutation of an ultralong DH with a severe codon bias toward mutation to cysteine. These unusual antibodies can be elicited to recognize defined antigens through the knob domain. Thus, the bovine immune system produces an antibody repertoire composed of ultralong CDR H3s that fold into a diversity of minidomains generated through combinations of somatically generated disulfides.

Wang F ，Ekiert DC ，Ahmad I ，Yu W ，Zhang Y ，Bazirgan O ，Torkamani A ，Raudsepp T ，Mwangi W ，Criscitiello MF ，Wilson IA ，Schultz PG ，Smider VV ... -  《-》

Immunogenetic factors driving formation of ultralong VH CDR3 in Bos taurus antibodies.

Deiss TC ，Vadnais M ，Wang F ，Chen PL ，Torkamani A ，Mwangi W ，Lefranc MP ，Criscitiello MF ，Smider VV ... -  《-》