Crystal Structure of the Pre-fusion Nipah Virus Fusion Glycoprotein Reveals a Novel Hexamer-of-Trimers Assembly
Fig 3
Formation of F hexamers-of-trimers in solution.
(A) 100μg of 50 mg/mL sF cross-linked with 0.08% glutaraldehyde was subjected to 10–25% sucrose gradient ultracentrifugation and fractionated. 10μL of each fraction was analyzed on Blue Native PAGE followed by western blotting. F oligomers were probed by monoclonal mouse anti-F specific antibody. The bottom and top gradient fractions are indicated. Arrows indicate higher-order oligomers formed from F trimers. (B) Hexameric sF assemblies imaged with negative stain EM of pooled fractions 1–6 from panel (A). It shows the cross-linked NiV-F particles boxed out from raw images. The majority of NiV-F oligomers appear as hexamers-of-trimers. (C) A small density slab (7.4 nm thick) from an electron tomogram of NiV-F decorated VLP showing the hexameric arrangement of the NiV-F spikes on the VLP. The individual NiV-F spikes are ~8–10 nm long with a ~1–2 nm stalk, consistent with the electron microscopy studies of soluble PIV5-F in its pre-fusion state [59]. The hexameric assembly revealed in NiV-sFGCNt crystal structure was shown next to it as reference.