Covalent dimerization of camelidae anti-human TNF-alpha single domain antibodies by the constant kappa light chain domain improves neutralizing activity.
The tumor necrosis factor-alpha (TNFalpha) plays an important role in a number of chronic inflammatory disorders. Monoclonal camelidae variable heavy chain domain-only antibodies (V(H)H) have been developed to antagonize the action of human TNFalpha (anti-TNF-V(H)H). Here we describe a strategy to obtain functional dimeric anti-TNF-V(H)H molecules, based on the C-terminal fusion of a kappa light chain domain to the anti-TNF-V(H)H. The resulting fusion protein was transiently expressed by use of viral vectors in Nicotiana benthamiana((Nb)) leaves and purified. Competitive ELISA and cell cytotoxicity assays revealed that the dimerized anti-(Nb)TNF-V(H)H(Ckappa) proteins blocked TNFalpha-activity more effectively than either the monomeric Escherichia coli((Ec)) produced anti-(Ec)TNF-V(H)H or the monomeric anti-(Nb)TNF-V(H)H(Ckappa). We suggest that enhanced inhibition shown by dimeric anti-(Nb)TNF-V(H)H(Ckappa) proteins is achieved by an increase in avidity.
Giersberg
Martin
M
Institute of Plant Genetics and Crop Plant Research (IPK), Phytoantibodies, D-06466 Gatersleben, Germany.
Floss
Doreen M
DM
Kipriyanov
Sergey
S
Conrad
Udo
U
Scheller
Jürgen
J
D000911:Antibodies, Monoclonal / Q000235:genetics / Q000494:pharmacology*; D057134:Antibodies, Neutralizing / Q000235:genetics / Q000494:pharmacology*; D019281:Dimerization; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D007147:Immunoglobulin Light Chains / Q000235:genetics / Q000494:pharmacology*; D007155:Immunologic Factors / Q000235:genetics / Q000494:pharmacology*; D010942:Plant Viruses / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000494:pharmacology; D014026:Nicotiana / Q000235:genetics; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors*
D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D007147:Immunoglobulin Light Chains; D007155:Immunologic Factors; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha