Thermolysin activation mutants with changes in the fusogenic region of an influenza virus hemagglutinin.

Influenza virus A/seal/Mass/1/80 (H7N7) mutants were obtained; the hemagglutinins (HAs) of the mutants were not activated by trypsin, as in the wild-type virus, but by thermolysin. The mutants grew efficiently under multiple replication cycle conditions and formed plaques in chicken embryo cells only when thermolysin was added to the culture medium. They exhibited hemolytic activity and induced protective immunity in chickens after an asymptomatic course of infection. Nucleotide sequencing of the HA gene and direct amino acid sequencing showed that insertion of a single leucine into the fusion peptide of the HA2 chain close to the cleavage site and a shift of the cleavage site toward the C terminus by one amino acid were responsible for the changes in the biological properties of the thermolysin activation mutants. Revertants could be obtained when trypsin or trypsin-like endoproteases were present in the virus-producing system.