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Abstract <p>COVID-19 is a rapidly disseminating disease whose causative agent is severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite the high investigation pace and the status of the most studied virus there are still blank spots in the understanding of SARS-CoV-2 mechanism of entry into the host cell. The most important role in this process is played by the viral spike (S) protein, particularly its transmembrane (TM) domain, whereon structural data are quite contradictory. In this work, we present a fast and high-performance method of cell-free expression and purification of the wild-type TM domain of the S protein including juxtamembrane parts. The purified peptide was incorporated into membrane mimicking dodecyl phosphocholine (DPC) micelles and was assessed as appropriate for further structural and dynamic investigation in nuclear magnetic resonance (NMR) experiments. We discuss the possible influence of amino acid sequence mutations, expression and purification methods.</p>

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Transmembrane Domain of Wild Type Spike Protein from SARS-CoV-2: Cell-Free Expression and Fast Purification for Structural-Dynamic NMR Studies

  • 期刊论文

  • 发表日期：

    2025年11月14日

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  • S. M. Sudareva,
  • Ya. V. Bershatsky,
  • A. S. Urban,
  • Yu. A. Zagryadskaya,
  • E. T. Aliper,
  • R. G. Efremov,
  • D. Zhu,
  • I. S. Okhrimenko,
  • E. V. Bocharov

摘要

Abstract

COVID-19 is a rapidly disseminating disease whose causative agent is severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite the high investigation pace and the status of the most studied virus there are still blank spots in the understanding of SARS-CoV-2 mechanism of entry into the host cell. The most important role in this process is played by the viral spike (S) protein, particularly its transmembrane (TM) domain, whereon structural data are quite contradictory. In this work, we present a fast and high-performance method of cell-free expression and purification of the wild-type TM domain of the S protein including juxtamembrane parts. The purified peptide was incorporated into membrane mimicking dodecyl phosphocholine (DPC) micelles and was assessed as appropriate for further structural and dynamic investigation in nuclear magnetic resonance (NMR) experiments. We discuss the possible influence of amino acid sequence mutations, expression and purification methods.