Isotope-edited nuclear magnetic resonance study of Fv fragment of anti-dansyl mouse monoclonal antibody: recognition of the dansyl hapten.

An isotope-edited proton nuclear magnetic resonance study is reported of Fv, which is the smallest antigen recognition unit composed of VH and VL domains. Fv has been obtained by clostripain digestion of a short-chain anti-dansyl mouse IgG2a monoclonal antibody Igarashi, T., Sato, M., Katsube, Y., Takio, K., Tanaka, T., Nakanishi, M., & Arata, Y. (1990) Biochemistry 29, 5727-5733]. A variety of stable-isotope-labeled anti-dansyl Fv analogues have been prepared. The aromatic proton resonances for all Tyr residues of the Fv fragment have been assigned in the absence and presence of epsilon-dansyl-L-lysine by means of isotope-edited homonuclear and heteronuclear two-dimensional NMR experiments. On the basis of the established assignments, it has been concluded that the dansyl ring is bound through Tyr-96H and Tyr-104H to both ends of H3, the third hypervariable region of the heavy chain. We also suggest that the antigen binding results in the formation of a hydrophobic core comprising the dansyl ring and the aromatic rings of Tyr-96H and Tyr-104H.