title: Camel single-domain antibodies as modular building units in bispecific and bivalent antibody constructs.
abstract: Single-domain antibodies against various antigens are isolated from the unique heavy-chain antibodies of immunized camels and llamas. These minimal sized binders are very robust and bind the antigen with high affinity in a monomeric state. We evaluated the feasibility to produce soluble, functional bispecific and bivalent antibodies in Escherichia coli with camel single-domain antibody fragments as building blocks. Two single-domain antibody fragments were tethered by the structural upper hinge of a natural antibody to generate bispecific molecules. This linker was chosen for its protease resistance in serum and its natural flexibility to reorient the upstream and downstream located domains. The expression levels, ease of purification, and the solubility of the recombinant proteins were comparable with those of the constituent monomers. The individual moieties fully retain the binding capacity and the binding characteristics within the recombinant bispecific constructs. The easy generation steps and the biophysical properties of these bispecific and bivalent constructs based on camel single-domain antibody fragments makes them particularly attractive for use in therapeutic or diagnostic programs.
authors.affiliation: Department of Ultrastructure, Vlaams Interuniversitair Instituut voor Biotechnologie, Vrije Universiteit Brussel, Paardenstraat 65, B-1640 Sint Genesius Rode, Belgium. kconrath@vub.ac.be
authors.forename: K
authors.initials: K
authors.lastname: Els Conrath
authors.forename: M
authors.initials: M
authors.lastname: Lauwereys
authors.forename: L
authors.initials: L
authors.lastname: Wyns
authors.forename: S
authors.initials: S
authors.lastname: Muyldermans
mesh_terms: D000595:Amino Acid Sequence; D000681:Amylases / Q000037:antagonists & inhibitors; D000818:Animals; D000906:Antibodies / Q000737:chemistry*; D018033:Antibodies, Bispecific / Q000737:chemistry*; D019921:Biotinylation; D015153:Blotting, Western; D002161:Camelids, New World; D002162:Camelus; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
chemical_list: D000906:Antibodies; D018033:Antibodies, Bispecific; D011994:Recombinant Proteins; D000681:Amylases