EBOV Glycoprotein Structural Characterization

T42V/T230V GP_33-632Δmuc was transiently expressed in HEK293T cells with an N-terminal haemaglutinin tag for purification via affinity chromatography. GP was natively deglycosylated using PNGaseF and complexed with a SeMet-containing KZ52 antibody, and the resulting trimeric GP-Fab complex was purified by size exclusion chromatography. Crystals were grown in 8.5% (w/v) PEG 10,000, 0.1 M Tris-HCl pH 8.5, 0.4 M sodium acetate and 10% (v/v) PEG 200 and diffracted to 3.4 Å resolution. Phases were obtained by combination of the Fab selenium anomalous signal and molecular replacement using an independently determined structure of the uncomplexed KZ52 Fab as a search model. Interpretable electron density maps with clear secondary structural elements and solvent boundaries were obtained after NCS-averaged density modification. The structure was refined to final R_work and R_free values of 26.2% and 30.2%, respectively. Atomic coordinates and structure factors for the reported crystal structure have been deposited with the Protein Data Bank under accession code 3CSY.