Alonso-Romero, J. | Search bioRxiv

bioRxiv2026

SARS-CoV-2 membrane protein biogenesis

Viral protein biogenesis underpins every viral life cycle stage, and elucidating these processes could reveal fundamental principles of virus-host interaction, and vulnerabilities amenable to therapeutic targeting. Here we apply biophysical, molecular, and cell biology techniques to investigate the insertion, folding, and oligomerization of the SARS-CoV-2 M protein. We describe the sequential co-translational insertion of the hydrophobic core, and demonstrate that the cytosolic C-terminal domain undergoes slower adoption of its tertiary structure. Additionally, we characterize how the transmembrane domain bundle facilitates M-protein oligomerization. Our results reveal a hydrophobic residue cluster that is essential for protein folding and co-translational dimerization. Additionally, we identify the cellular machinery responsible for targeting and inserting the M protein into the ER membrane, and chaperones and cofactors that may contribute to proper folding.

molecular biology↗

bioRxiv2024

The sequence and structural integrity of the SARS-CoV-2 Spike protein transmembrane domain is crucial for viral entry

The Spike (S) protein of SARS-CoV-2 is a type I membrane protein that mediates target cell recognition and membrane fusion. While its transmembrane domain (TMD) is traditionally viewed as a passive anchor to the viral envelope, emerging evidence suggests that TMDs often play active roles in the biogenesis and function of membrane proteins. Here, we investigated the functional role of the SARS-CoV-2 S protein TMD during viral entry. To this end, we introduced a series of amino acid substitutions and insertions within the hydrophobic core of the TMD and assessed their impact on S protein activity. Our findings reveal that the SARS-CoV-2 S protein is susceptible to alterations in its TMD. Functional determinants, including sequence features and structural parameters critical for viral entry, are distributed throughout the TMD, with a more pronounced contribution from its N-terminal region. We also demonstrate that the relative orientation of the regions flanking the TMD influences viral entry. Finally, our data suggest that the TMD mediates homo-oligomerization through a motif enriched in small residues, underscoring its functional importance beyond membrane anchoring.

molecular biology↗