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bioRxiv · 10.1101/2024.03.07.583986

A tale of two fusion proteins: understanding the metastability of human respiratory syncytial virus and metapneumovirus and implications for rational design of uncleaved prefusion-closed trimers

Lee, Y.-Z.·Han, J.·Zhang, Y.-N.·Ward, G.·Gomes, K. B.·Auclair, S.·Stanfield, R. L.·He, L.·Wilson, I. A.·Zhu, J.
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Abstract

Respiratory syncytial virus (RSV) and human metapneumovirus (hMPV) cause human respiratory diseases and are major targets for vaccine development. In this study, we designed uncleaved prefusion-closed (UFC) trimers for the fusion (F) proteins of both viruses by examining mutations critical to F metastability. For RSV, we assessed four previous prefusion F designs, including the first and second generations of DS-Cav1, SC-TM, and 847A. We then identified key mutations that can maintain prefusion F in a native-like, closed trimeric form (up to 76%) without introducing any interprotomer disulfide bond. For hMPV, we developed a stable UFC trimer with a truncated F2-F1 linkage and an interprotomer disulfide bond. Tens of UFC constructs were characterized by negative-stain electron microscopy (nsEM), x-ray crystallography (11 RSV-F and one hMPV-F structures), and antigenic profiling. Using an optimized RSV-F UFC trimer as bait, we identified three potent RSV neutralizing antibodies (NAbs) from a phage-displayed human antibody library, with a public NAb lineage targeting sites O and V and two cross-pneumovirus NAbs recognizing site III. In mouse immunization, rationally designed RSV-F and hMPV-F UFC trimers induced robust antibody responses with high neutralizing titers. Our study provides a foundation for future prefusion F-based RSV and hMPV vaccine development. ONE-SENTENCE SUMMARYThe metastability analysis of fusion proteins has informed rational design of uncleaved prefusion-closed trimers for RSV and hMPV vaccine development.

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Lee, Y.-Z., Han, J., Zhang, Y.-N., Ward, G., Gomes, K. B., Auclair, S., Stanfield, R. L., He, L., Wilson, I. A., Zhu, J.. 2024-03-08. A tale of two fusion proteins: understanding the metastability of human respiratory syncytial virus and metapneumovirus and implications for rational design of uncleaved prefusion-closed trimers. https://doi.org/10.1101/2024.03.07.583986

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