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bioRxiv · 10.1101/2023.09.30.560318

Synthesis, Insertion and Characterization of SARS-CoV-2 Membrane Protein Within Lipid Bilayers

Zhang, Y.·Anbir, S.·Mctiernan, J.·Li, S.·Worcester, M.·Mishra, P.·Colvin, M. E.·Gopinathan, A.·Mohideen, U.·Zandi, R.·Kuhlman, T. E.
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Abstract

SUMMARY/ABSTRACTThe membrane protein (M) is the most abundant structural protein in the SARS-CoV-2 virus and functions exclusively as a membrane-embedded homodimer. M protein is required for the formation of the SARS-CoV-2 virus particle and has been shown to interact with the Spike and Envelope proteins, as well as the RNA-packaging Nucleocapsid protein. Our knowledge of M protein is very limited due to its small size and challenges in expressing enough protein for use in structural and biophysical experiments. We report the successful development of a SUMO tag-based expression system to produce and purify significant quantities of M protein, and a method to insert the synthesized dimers into a suspended lipid membrane in a homogeneous orientation. We used AFM and Cryo-EM to image individual membrane-bound M protein dimers and characterize the configurations that they can assume. Our experimental results are in agreement with our molecular dynamics simulations which predict thinning of the membrane around the M protein and a propensity to induce local membrane curvature. Taken together, our results shed new light on M protein properties within the lipid bilayer and suggest mechanisms that could contribute to viral assembly and budding.

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Zhang, Y., Anbir, S., Mctiernan, J., Li, S., Worcester, M., Mishra, P., Colvin, M. E., Gopinathan, A., Mohideen, U., Zandi, R., Kuhlman, T. E.. 2023-10-02. Synthesis, Insertion and Characterization of SARS-CoV-2 Membrane Protein Within Lipid Bilayers. https://doi.org/10.1101/2023.09.30.560318

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