Fig. 5

Structural evidence for CR9501-enhanced opening of prefusion F trimers. a Model illustrating the strategy used to generate trimeric prefusion F bound simultaneously by CR9501 and motavizumab, showing prefusion F protomers in blue, light gray and dark gray, CR9501 in pink and light pink, and motavizumab in orange and tan. b Cartoon depicts the biological RSV F trimer (left) and the molecular replacement solution obtained in space group P6₁ (right), shown without Fabs for clarity. Prefusion F protomers are colored blue, light gray, and dark gray. c The molecular replacement solution from the crystal structure of prefusion F bound to CR9501 and motavizumab, with six copies of the asymmetric unit shown and the sixfold screw axis indicated in the center (top). Prefusion F is shown as blue ribbons and motavizumab and CR9501 Fabs are shown as molecular surfaces. CR9501 heavy and light chains are colored dark pink and light pink and the motavizumab Fab heavy and light chains are colored orange and tan, respectively. The Foldon motif was modeled on to the C-terminus of each monomer, using a previously determined prefusion F structure in which this domain is visible (PDB ID: 4MMV) (bottom). d Negative-stain EM of the prefusion F-motavizumab complex showed primarily top-down views of the closed, trimeric complex (scale bar = 50 nm). A representative class average shows a closed trimer with three Fabs bound and the cartoon illustrates the orientation viewed in the class average. e Negative-stain EM of the prefusion F-CR9501 complex revealed multiple particle classes. Three representative class averages show different orientations of the closed, trimeric complex, whereas the fourth comprised individual monomers within splayed-open trimers. The inset shows a subset of the splayed-open trimers used to generate the fourth class average (right) and the cartoons illustrate the orientation of the splayed-open trimer thought to be present in the particles shown