Using monoclonal antibodies to characterize a sequential epitope on the group I allergen of Bermuda grass pollen.

Abstract

Cyn d 1, the group I allergen of Bermuda grass pollen, had been purified and characterized. A sequential B cell epitope on Cyn d 1 was studied with monoclonal antibodies (MoAbs). Cyn d 1 was cleaved by Achromobacter protease I into fragments, and the resulting peptides were fractionated on reversed-phase columns before being reacted with anti-Cyn d 1 MoAbs in a radioimmunoassay. A Cyn d 1 fragment recognized by its MoAb was selected for Edman degradation. A synthetic peptide was constructed according to the determined sequence. The epitope on Cyn d 1 recognized by MoAb 18-53 was found to be conformation independent, since its activity was not changed after sodium periodate, guanidine or urea treatment. The enzyme-cleaved fragment containing this epitope was determined to be DVDKPPFDGMTACGNEPIF which corresponds to the N-terminal 46-64 residues of Cyn d 1. The presence of this sequence in the epitope recognized by MoAb 18-53 was demonstrated by enzyme immunoassay and further confirmed by inhibition of binding enzyme immunoassay with synthetic peptides. Some cross-reactivity with the N-terminal 45-63 residues of Lol p 1 was also found. The primary structure of a sequential epitope on Cyn d 1 was determined, and its activity was confirmed with peptides synthesized according to the determined sequence.