MOLECULAR AND STRUCTURAL-ANALYSIS OF A CONTINUOUS BIRCH PROFILIN EPITOPE DEFINED BY A MONOCLONAL-ANTIBODY

The interaction of a mouse monoclonal antibody (4A6) and birch profili n, a structurally well conserved actin and phosphoinositide-binding pr otein and cross-reactive allergen, was characterized, In contrast to s erum IgE from allergic patients, which shows cross-reactivity with mos t plants, monoclonal antibody 4A6 selectively reacted with tree pollen profilins. Using synthetic overlapping peptides, a continuous hexapep tide epitope was identified. The exchange of a single amino acid (Gln- 47 --> Glu) within the epitope was found to abolish the bind ing of mo noclonal antibody 4A6 to other plant profilins. The MMR analyses of th e birch and the nonreactive timothy grass profilin peptides showed tha t the loss of binding was not due to major structural differences. Bot h peptides adopted extended conformations similar to that observed for the epitope in the x-ray crystal structure of the native birch profil in. Binding studies with peptides and birch profilin mutants generated by in vitro mutagenesis demonstrated that the change of Gln-47 to aci dic amino acids (e.g. Glu or Asp) led to electrostatic repulsion of mo noclonal antibody 4A6. In conclusion the molecular and structural anal yses of the interaction of a monoclonal antibody with a continuous pep tide epitope, recognized in a conformation similar to that displayed o n the native protein, are presented.