Structure of SARS-CoV-2 M protein in lipid nanodiscs

Abstract

SARS-CoV-2 encodes four structural proteins incorporated into virions, spike (S), envelope (E), nucleocapsid (N), and membrane (M). M plays an essential role in viral assembly by organizing other structural proteins through physical interactions and directing them to sites of viral budding. As the most abundant protein in the viral envelope and a target of patient antibodies, M is a compelling target for vaccines and therapeutics. Still, the structure of M and molecular basis for its role in virion formation are unknown. Here, we present the cryo-EM structure of SARS-CoV-2 M in lipid nanodiscs to 3.5 Å resolution. M forms a 50 kDa homodimer that is structurally related to the SARS-CoV-2 ORF3a viroporin, suggesting a shared ancestral origin. Structural comparisons reveal how intersubunit gaps create a small, enclosed pocket in M and large open cavity in ORF3a, consistent with a structural role and ion channel activity, respectively. M displays a strikingly electropositive cytosolic surface that may be important for interactions with N, S, and viral RNA. Molecular dynamics simulations show a high degree of structural rigidity in a simple lipid bilayer and support a role for M homodimers in scaffolding viral assembly. Together, these results provide insight into roles for M in coronavirus assembly and structure.

Systematic Exploration of SARS-CoV-2 Adaptation to Vero E6, Vero E6/TMPRSS2, and Calu-3 Cells

“… Positions of the focus adaptation mutations on the proteins: ( a ) spike glycoprotein, ( b ) nucleocapsid, ( c ) NSP1, ( d ) membrane glycoprotein, and ( e ) NSP3 PLpro. The AlphaFold2 ( Jumper et al 2021 ; Mirdita et al 2022 ) models or Protein Data Bank structures 6vsb ( Hwang et al 2020 ), 8ctk ( Dolan et al 2022 ), or 7cjd ( Gao et al 2021 ) are drawn in cartoon representation with gray transparent surface. Positions harboring focused adaptation mutations ( fig.…”

“…For the trimeric spike glycoprotein, the predicted structure of the domains surrounding the missing loops were superimposed to those of the cryogenic electron microscopy (cryo-EM) structure (Protein Data Bank (PDB) number: 6vsb; Hwang et al 2020 ), and the predicted coordinates of the loops were added to the experimental structure without further optimization. The cryo-EM structure of the spike glycoprotein (PDB number: 6vsb; Hwang et al 2020 ), and the membrane glycoprotein in a lipid nanodisc (PDB number: 8ctk; Dolan et al 2022 ), as well as the crystal structure of NSP3 PLpro (PDB 7cjd; Gao et al 2021 ), were obtained from the Protein Data Bank maintained by Research Collaboratory for Structural Bioinformatics ( https://www.rcsb.org/ ; Berman et al 2000 ).…”

Perturbation of the host cell Ca2+ homeostasis and ER-mitochondria contact sites by the SARS-CoV-2 structural proteins E and M

“…The cryo-EM structure of the SARS-CoV2 M protein has been recently solved in two different conformations. M protein forms a dimer [ 41 ] but can also further assemble into higher-order oligomers [ 42 ]. Although their specific role is still largely unclear, M/E, M/N or M/N/E interactions are important for efficient virion assembly [ 43 ].…”

Accelerating antiviral drug discovery: lessons from COVID-19

“…The availability of meaningful functional assays and linked structural data can greatly facilitate the development of chemical probes for previously untargeted viral proteins. In particular, advances in cryo-electron microscopy now allow elucidation of complex protein structures 14 – 16 . Although many chemical probes have been developed for human targets, fewer probes and structural data are available for viral targets.…”