title: High-affinity lamprey VLRA and VLRB monoclonal antibodies.
abstract: Lamprey are members of the ancestral vertebrate taxon (jawless fish), which evolved rearranging antigen receptors convergently with the jawed vertebrates. But instead of Ig superfamily domains, lamprey variable lymphocyte receptors (VLRs) consist of highly diverse leucine-rich repeats. Although VLRs represent the only known adaptive immune system not based on Ig, little is known about their antigen-binding properties. Here we report robust plasma VLRB responses of lamprey immunized with hen egg lysozyme and beta-galactosidase (beta-gal), demonstrating adaptive immune responses against soluble antigens. To isolate monoclonal VLRs, we constructed large VLR libraries from antigen-stimulated and naïve animals in a novel yeast surface-display vector, with the VLR C-terminally fused to the yeast Flo1p surface anchor. We cloned VLRB binders of lysozyme, beta-gal, cholera toxin subunit B, R-phycoerythrin, and B-trisaccharide antigen, with dissociation constants up to the single-digit picomolar range, equivalent to those of high-affinity IgG antibodies. We also isolated from a single lamprey 13 anti-lysozyme VLRA clones with affinities ranging from low nanomolar to mid-picomolar. All of these VLRA clones were closely related in sequence, differing at only 15 variable codon positions along the 244-residue VLR diversity region, which augmented antigen-binding affinity up to 100-fold. Thus, VLRs can provide a protective humoral antipathogen shield. Furthermore, the broad range of nominal antigens that VLRs can specifically bind, and the affinities achieved, indicate a functional parallelism between LRR-based and Ig-based antibodies. VLRs may be useful natural single-chain alternatives to conventional antibodies for biotechnology applications.
authors.affiliation: Center of Marine Biotechnology, University of Maryland Biotechnology Institute, Baltimore, MD 21202, USA.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tasumi
authors.forename: C Alejandro
authors.initials: CA
authors.lastname: Velikovsky
authors.forename: Gang
authors.initials: G
authors.lastname: Xu
authors.forename: S Annie
authors.initials: SA
authors.lastname: Gai
authors.forename: K Dane
authors.initials: KD
authors.lastname: Wittrup
authors.forename: Martin F
authors.initials: MF
authors.lastname: Flajnik
authors.forename: Roy A
authors.initials: RA
authors.lastname: Mariuzza
authors.forename: Zeev
authors.initials: Z
authors.lastname: Pancer
mesh_terms: D000222:Adaptation, Physiological; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000915:Antibody Affinity; D001483:Base Sequence; D007798:Lampreys / Q000276:immunology*; D000090263:Leucine-Rich Repeat Proteins; D008969:Molecular Sequence Data; D011506:Proteins / Q000276:immunology*; D011946:Receptors, Antigen / Q000276:immunology* / Q000502:physiology
chemical_list: D000911:Antibodies, Monoclonal; D000090263:Leucine-Rich Repeat Proteins; D011506:Proteins; D011946:Receptors, Antigen