The SARS-CoV-2 spike is the primary target of virus-neutralizing antibodies and critical to the development of effective vaccines against COVID-19. Here, we demonstrate that the prefusion-stabilized two-proline “S2P” spike—widely employed for laboratory work and clinical studies—unfolds when stored at 4 °C, physiological pH, as observed by electron microscopy (EM) and differential scanning calorimetry, but that its trimeric, native-like conformation can be reacquired by low pH treatment. When stored for approximately 1 week, this unfolding does not significantly alter antigenic characteristics; however, longer storage diminishes antibody binding, and month-old spike elicits virtually no neutralization in mice despite inducing high ELISA-binding titers. Cryo-EM structures reveal the folded fraction of spike to decrease with aging; however, its structure remains largely similar, although with varying mobility of the receptor-binding domain. Thus, the SARS-CoV-2 spike is susceptible to unfolding, which affects immunogenicity, highlighting the need to monitor its integrity.
【저자키워드】 COVID-19 vaccine, RBD, receptor-binding domain, SARS-CoV-2 spike, NTD, N-terminal domain, PBS, phosphate buffered saline, CTF, contrast transfer function, DSC, differential scanning calorimetry, pH-induced folding, S2P, unfolding, BLI, bio-layer interferometry, cryo-EM, cryo-electron microscopy, EM, electron microscopy, NS-EM, negative-stain electron microscopy, 【초록키워드】 COVID-19, Treatment, Structure, neutralization, Laboratory, Antibody binding, mice, Microscopy, Critical, physiological, antigenic, fraction, MONITOR, Affect, Alter, susceptible, decrease, virus-neutralizing antibody, trimeric, significantly, elicit, the receptor-binding domain, effective vaccine, diminishe, highlighting, the SARS-CoV-2, 【제목키워드】 SARS-CoV-2, age,