Chromatographic isolation of the hemagglutinin polypeptides from influenza virus vaccine and determination of their amino-terminal sequences.

Abstract

The influenza virus hemagglutinin polypeptides HA1 and HA2 were purified by gel filtration in the presence of sodium dodecyl sulfate from a vaccine preparation of the recombinant strain Heq1N2. Use of this technique for purification of the hemagglutinin polypeptides eliminated the need for proteolytic agents for removal of the hemagglutinin from the virus particles; 100-300 mg of virus yielded 10-30 mg viral protein/chromatographic cycle. Because proteolysis is not required to remove the spikes from the viral envelope, the envelope-embedded HA2 polypeptide was purified in its entirety for structural analysis. Amino-terminal sequence analysis of the smaller polypeptide, HA2, revealed a cyclic repetition of glycyl residues through the first 24 residues at every 3rd-4th position. The sequence through the first 10 residues was identical to that presented for other type A influenza viruses. The HA1 (Heq1) polypeptide had different amino acids at 3 or 4 of the first 10 residues of the amino-terminal sequence when compared to HA1 from H0, H1 or H2 subtypes. This study demonstrates the feasibility of the use of vaccine virus as a source of large quantities of viral protein for determination of primary structure.