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bioRxiv · 10.1101/2020.07.10.197814

Stabilizing the Closed SARS-CoV-2 Spike Trimer

Jarek Juraszek·Lucy Rutten·Sven Blokland·Pascale Bouchier·Richard Voorzaat·Tina Ritschel·Mark J.G. Bakkers·Ludovic L.R. Renault·Johannes P.M. Langedijk
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Abstract

The trimeric spike (S) protein of SARS-CoV-2 is the primary focus of most vaccine design and development efforts. Due to intrinsic instability typical of class I fusion proteins, S tends to prematurely refold to the post-fusion conformation, compromising immunogenic properties and prefusion trimer yields. To support ongoing vaccine development efforts, we report the structure-based design of soluble S trimers, with increased yields and stabilities, based on introduction of single point mutations and disulfide-bridges. We identify two regions in the S-protein critical for the proteins stability: the heptad repeat region 1 of the S2 subunit and subunit domain 1 at the interface with S2. We combined a minimal selection of mostly interprotomeric mutations to create a stable S-closed variant with a 6.4-fold higher expression than the parental construct while no longer containing a heterologous trimerization domain. The cryo-EM structure reveals a correctly folded, predominantly closed pre-fusion conformation. Highly stable and well producing S protein and the increased understanding of S protein structure will support vaccine development and serological diagnostics.

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Jarek Juraszek, Lucy Rutten, Sven Blokland, Pascale Bouchier, Richard Voorzaat, Tina Ritschel, Mark J.G. Bakkers, Ludovic L.R. Renault, Johannes P.M. Langedijk. 2020-07-10. Stabilizing the Closed SARS-CoV-2 Spike Trimer. https://doi.org/10.1101/2020.07.10.197814

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