Evolutionarily conserved residues at an oligomerization interface of the influenza A virus neuraminidase are essential for viral survival

•Twenty-one amino acids at NA were identified as crucial residues for viral survival.•Lack of NA enzymatic activity is not a sole factor in influenza viral lethality.•Ala substitution at the NA oligomerization interface disrupts the stability of the NA dimer/tetramer.•The oligomerization of NA is important for viral viability.

Neuraminidase (NA) is a homotetramer viral surface glycoprotein that is essential for virus release during influenza virus infections. Previous studies have not explored why influenza NA forms a tetramer when the bacterial monomer NA already exhibits excellent NA enzymatic activity levels. In this study, we focused on 28 highly conserved residues among all NA subtypes, identifying 21 of 28 positions as crucial residues for viral survival by using reverse genetics. Maintaining NA enzymatic activity levels is critical and numerous conserved residues were located at the oligomerization interface; however, these mutations did not affect NA enzymatic activity levels or NA cellular localization, but rather affected the stability of NA oligomerization, suggesting that the oligomerization of NA is essential for viral viability. An increased understanding of the biological functions of NA, in particular NA oligomerization, could facilitate an alternative design for antivirals to combat influenza virus infections.