Crystal structure of a human single domain antibody dimer formed through V(H)-V(H) non-covalent interactions

Single-domain antibodies (sdAbs) derived from human V_H are considered to be less soluble and prone to aggregate which makes it difficult to determine the crystal structures. In this study, we isolated and characterized two anti-human epidermal growth factor receptor-2 (HER2) sdAbs, Gr3 and Gr6, from a synthetic human V_H phage display library. Size exclusion chromatography and surface plasmon resonance analyses demonstrated that Gr3 is a monomer, but that Gr6 is a strict dimer. To understand this different molecular behavior, we solved the crystal structure of Gr6 to 1.6 Å resolution. The crystal structure revealed that the homodimer assembly of Gr6 closely mimics the V_H-V_L heterodimer of immunoglobulin variable domains and the dimerization interface is dominated by hydrophobic interactions.