Antibodies provide the first line of defence against invading organisms. Neutralizing antibodies usually react with surface antigens of virus particles and it is therefore crucial to know the three-dimensional structure of the virus protein and its complex with antibodies in order to understand the mechanism by which antibodies inhibit the functions of the protein. Currently the three-dimensional structure of only one virus protein (type A influenza virus neuraminidase) complexed with antibody Fab fragments has been solved by X-ray crystallography (Colman et al, 1987b). The neuraminidases from influenza virus type A and B differ in amino acid sequence by 75% and are antigenically unrelated. The objective of this proposal is to determine the three-dimensional structure of type B influenza virus neuraminidase complexed with antibody Fab fragments by X-ray diffraction analysis of complex crystals at high resolution. Crystals of B/Lee/40 NA complexed with monoclonal antibody Fab fragments which diffract X-rays to 3.0 angstrom resolution have already been grown. The following specific aims are proposed: 1. To collect and process a complete data set at 3.0 angstrom resolution using native complex crystals. The data will be collected at synchrotron radiation sources by oscillation protography. Film processing will be done with a supercomputer and programs developed at PURDUE University. 2. To determine the non- crystallographic symmetry using Rossmann's rotation function and to try to determine the phases by the molecular replacement method using known structures of Fab fragments and type A neuraminidase. 3. To determine the phases by the isomorphous replacement method using heavy atom derivatives, if the molecular replacement method using structural models is unsuccessful. Molecular replacement techniques will then be used to improve, or even to determine, the high resolution phases. 4. To build the atomic model according to the electron density map and refine the structure by the Hendrickson and Konnert restrained least-squares refinement procedure. These experiments will define precisely the structure of an epitope on type B influenza virus neuraminidase and the corresponding binding site on the antibody molecule.
| Luo, M; Laver, W G; Air, G (1992) Unusual diffraction of type B influenza virus neuraminidase crystals. Acta Crystallogr A 48 ( Pt 3):263-6 |
| Lin, Y J; Luo, M; Laver, W G et al. (1990) New crystalline forms of neuraminidase of type B human influenza virus. J Mol Biol 214:639-40 |
