The Chemistry of Antigenic Variation in Influenza a Virus Hemagglutinin

Summary

Single-step antigenic mutants, derived experimentally from the strain A/NT60/68 (H3N2), have been used to study the chemistry of antigenic variation within the present subtype of influenza A virus. These mutants mimic the antigenic drift of field strains that arose naturally between 1968 and 1973. Hemagglutinin, the major viral coat antigen, was prepared free of neuraminidase by controlled digestion of each virus strain with the protease, bromelain. The antigenic reactivity of the hemagglutinin was not altered by this treatment.

The hemagglutinin preparations, after reduction and S-alkylation, were digested with trypsin and compared by fluorescent dansyl-peptide mapping on thin layers of silica gel. This sensitive mapping technique, operating on a nanomole of protein, permitted resolution of virtually all the tryptic peptides.

The maps of all strains were identical, except for one or two peptides migrating differently. The variable peptides were confined to the larger subunit (HAI) of the hemagglutinin molecule. Thus, variation within a hierarchic series of antigenic mutants is associated with minor changes in the primary structure of the larger hemagglutinin subunit. Since the strains examined differed only in their antigenic properties, it is likely that the variable peptide in which amino acid substitutions occurred formed part of the antigenic determinant.