A novel method to identify spike stabilizing mutations for COVID-19 vaccine development Coronavirus Disease COVID Vaccine SARSCoV2 UofIllinois scrippsresearch CWRUSOM
By Neha MathurSep 29 2022Reviewed by Benedette Cuffari, M.Sc. In a recent study published on the bioRxiv* preprint server, researchers discuss a robust method to identify mutations that simultaneously improve the expression and stabilize the prefusion conformation of the severe acute respiratory syndrome coronavirus 2 spike glycoprotein.
Previous studies have shown that several other mutations could also improve SARS-CoV-2 S expression. For example, an S construct known as HexaPro containing F817P, A892P, A899P, and A942P mutations, in addition to K986P and V987P, has been shown to prove S expression. A stable cell line that expresses human angiotensin-converting enzyme 2 and mNG21-10, which was named hACE2-expressing cells, was also developed.
Lastly, the researchers combined the validated fusion-incompetent mutations K986P, V987P, D994Q 181, and Q1005R to generate double, triple, and quadruple mutants of the membrane-bound or full-length S protein. The expression and fusion scores distribution of silent mutations were significantly different from those of nonsense mutations, thereby indicating that DMS experiments could distinguish mutants with different expression and fusogenicity levels.
In addition to K986P and V987P, the study method identified other mutations in HR1 and CH which had low adjusted fusion and high expression scores, of which included T961F, D994E, D994Q, and Q1005R. Notably, D994E and D994Q clustered at the same amino acid residue positions and were chemically similar.
Most SARS-CoV-2 neutralizing antibodies target the receptor-binding domain region of the S protein; therefore, HR1 and CH may be under low selection pressure. Another explanation for this observation is that other evolutionary constraints on HR1 and CH might be present in vivo.
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