Contacts between influenza virus N9 neuraminidase and monoclonal antibody NC10.
We are interested in studying how influenza virus escapes antibody inhibition. Based on the structure of the complex between N9 NA and monoclonal antibody NC10 Fab (R. L. Malby, W. R. Tulip, V. R. Harley, J. L. McKimm-Breschkin, W. G. Laver, R. G. Webster, and P. M. Colman, 1994, Structure 2, 733-746), we investigated the contribution made by individual amino acids to the stability of the complex. We made conservative changes in residues that are centrally located in the epitope and more drastic changes in peripheral contacts. The mutations made were N200L (removing an N-linked oligosaccharide), N329Q, N345Q, S370T, S372A, N400L, and K432M. Binding of each mutant to NC10 was quantitated by NA inhibition assays and ELISA. Except for N200L and N329Q, the mutants were inhibited by NC10 to the same extent as wild-type NA although with less affinity. The enzyme activity (K(cat)) of N200L is 80% reduced, indicating a defect in folding or assembly; therefore, the loss in binding activity due to the missing sugar residue cannot be assessed. The K(d) for N329Q is sixfold higher than for wild-type NA in the inhibition test, but the same as wild-type in ELISA, indicating a change in disposition of the antibody but no loss of affinity. The results show that the NC10 epitope can accommodate a change at any site and is not dominated by a few high-energy interactions as was found in the NC41 epitope. We propose that the difference lies in the contribution of buried water molecules to the NA-NC10 complex.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D000914:Antibodies, Viral / Q000737:chemistry / Q000276:immunology*; D000936:Antigen-Antibody Complex / Q000737:chemistry*; D000937:Antigen-Antibody Reactions; D000939:Epitopes; D006367:HeLa Cells; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000276:immunology*; D009980:Influenza A virus / Q000276:immunology*; D008961:Models, Structural; D016296:Mutagenesis; D009439:Neuraminidase / Q000737:chemistry / Q000276:immunology*; D011817:Rabbits; D014867:Water
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000936:Antigen-Antibody Complex; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D014867:Water; D009439:Neuraminidase