Li, X. Y. | bioRxiv Science

bioRxiv2022

A mosaic-type trimeric RBD-based COVID-19 vaccine candidate induces potent neutralization against Omicron and other SARS-CoV-2 variants

Large-scale populations in the world have been vaccinated with COVID-19 vaccines, however, breakthrough infections of SARS-CoV-2 are still growing rapidly due to the emergence of immune-evasive variants, especially Omicron. It is urgent to develop effective broad-spectrum vaccines to better control the pandemic of these variants. Here, we present a mosaic-type trimeric form of spike receptor-binding domain (mos-tri-RBD) as a broad-spectrum vaccine candidate, which carries the key mutations from Omicron and other circulating variants. Tests in rats showed that the designed mos-tri-RBD, whether used alone or as a booster shot, elicited potent cross-neutralizing antibodies against not only Omicron but also other immune-evasive variants. Neutralizing antibody titers induced by mos-tri-RBD were substantially higher than those elicited by homo-tri-RBD (containing homologous RBDs from prototype strain) or the inactivated vaccine BBIBP-CorV. Our study indicates that mos-tri-RBD is highly immunogenic, which may serve as a broad-spectrum vaccine candidate in combating SARS-CoV-2 variants including Omicron.

immunology↗

bioRxiv2020

In-cell destabilization of a homo-dimeric protein complex detected by DEER spectroscopy

The complexity of the cellular medium can affect proteins properties and therefore in-cell characterization of proteins is essential. We explored the stability and conformation of BIR1, the first baculoviral IAP repeat domain of X-chromosome-linked inhibitor of apoptosis (XIAP), as a model for a homo-dimer protein in human HeLa cells. We employed double electron-electron resonance (DEER) spectroscopy and labeling with redox stable and rigid Gd3+ spin labels at three protein residues, C12 (flexible region), E22C and N28C (part of helical residues 26-31) in the N-terminal region. In contrast to predictions by excluded volume crowding theory, the dimer-monomer dissociation constant KD was markedly higher in cells than in solution and dilute cell lysate. As expected, this increase was recapitulated under conditions of high salt concentrations given that a conserved salt bridge at the dimer interface is critically required for association. Unexpectedly, however, also the addition of a crowding agent such as Ficoll destabilized the dimer, suggesting that Ficoll forms specific interactions with the monomeric protein. Changes in DEER distance distributions were observed for the E22C site, which displayed reduced conformational freedom in cells. Although overall DEER behaviors at E22C and N28C were compatible with a predicted compaction of disordered protein regions by excluded volume effects, we were unable to reproduce E22C properties in artificially crowded solutions. These results highlight the importance of in-cell DEER measurements to appreciate the complexities of cellular in vivo effects on protein structures and functions.

biophysics↗