Structure-based design of prefusion-stabilized SARS-CoV-2 spikes

• Published in: Science (American Association for the Advancement of Science) Vol. 369; no. 6510; pp. 1501 - 1505
• Main Authors: Hsieh, Ching-Lin, Goldsmith, Jory A., Schaub, Jeffrey M., DiVenere, Andrea M., Kuo, Hung-Che, Javanmardi, Kamyab, Le, Kevin C., Wrapp, Daniel, Lee, Alison G., Liu, Yutong, Chou, Chia-Wei, Byrne, Patrick O., Hjorth, Christy K., Johnson, Nicole V., Ludes-Meyers, John, Nguyen, Annalee W., Park, Juyeon, Wang, Nianshuang, Amengor, Dzifa, Lavinder, Jason J., Ippolito, Gregory C., Maynard, Jennifer A., Finkelstein, Ilya J., McLellan, Jason S.
• Format: Journal Article
• Language: English
• Published: United States The American Association for the Advancement of Science 18.09.2020; American Association for the Advancement of Science
• Subjects: Amino Acid Substitution; Antibodies; Betacoronavirus - chemistry; Biochem; Coronaviridae; Coronavirus Infections - prevention & control; Coronaviruses; COVID-19; COVID-19 Vaccines; Cryoelectron Microscopy; Freeze thaw cycles; Freeze-thaw durability; Freeze-thawing; Freezing; Heat stress; Heat tolerance; Humans; Pandemics; Proline; Proline - chemistry; Protein Domains; Protein Stability; Proteins; Respiratory diseases; Room temperature; SARS-CoV-2; Severe acute respiratory syndrome coronavirus 2; Spike Glycoprotein, Coronavirus - chemistry; Vaccines; Viral diseases; Viral Vaccines - chemistry; Virology
• ISSN: 0036-8075; 1095-9203; 1095-9203
• DOI: 10.1126/science.abd0826

The development of therapeutic antibodies and vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is focused on the spike (S) protein that decorates the viral surface. A version of the spike ectodomain that includes two proline substitutions (S-2P) and stabilizes the prefusion conformation has been used to determine high-resolution structures. However, even S-2P is unstable and difficult to produce in mammalian cells. Hsieh et al. characterized many individual and combined structure-guided substitutions and identified a variant, named HexaPro, that retains the prefusion conformation but shows higher expression than S-2P and can also withstand heating and freezing. This version of the protein is likely to be useful in the development of vaccines and diagnostics. Science , this issue p. 1501 The design of stabilizing mutations in the SARS-CoV-2 spike protein allows for high-yield production of a critical vaccine antigen. The coronavirus disease 2019 (COVID-19) pandemic has led to accelerated efforts to develop therapeutics and vaccines. A key target of these efforts is the spike (S) protein, which is metastable and difficult to produce recombinantly. We characterized 100 structure-guided spike designs and identified 26 individual substitutions that increased protein yields and stability. Testing combinations of beneficial substitutions resulted in the identification of HexaPro, a variant with six beneficial proline substitutions exhibiting higher expression than its parental construct (by a factor of 10) as well as the ability to withstand heat stress, storage at room temperature, and three freeze-thaw cycles. A cryo–electron microscopy structure of HexaPro at a resolution of 3.2 angstroms confirmed that it retains the prefusion spike conformation. High-yield production of a stabilized prefusion spike protein will accelerate the development of vaccines and serological diagnostics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).