- Protocol for structure determination of SARS-CoV-2 main protease at near-physiological-temperature by serial femtosecond crystallography.
Protocol for structure determination of SARS-CoV-2 main protease at near-physiological-temperature by serial femtosecond crystallography.
STAR protocols (2022-02-24)
Fatma Betul Ertem, Omur Guven, Cengizhan Buyukdag, Oktay Gocenler, Esra Ayan, Busra Yuksel, Mehmet Gul, Gozde Usta, Barıs Cakılkaya, J Austin Johnson, E Han Dao, Zhen Su, Frederic Poitevin, Chun Hong Yoon, Christopher Kupitz, Brandon Hayes, Mengning Liang, Mark S Hunter, Alexander Batyuk, Raymond G Sierra, Gihan Ketawala, Sabine Botha, Çağdaş Dağ, Hasan DeMirci
PMID35194584
ABSTRACT
The SARS-CoV-2 main protease of (Mpro) is an important target for SARS-CoV-2 related drug repurposing and development studies. Here, we describe the steps for structural characterization of SARS-CoV-2 Mpro, starting from plasmid preparation and protein purification. We detail the steps for crystallization using the sitting drop, microbatch (under oil) approach. Finally, we cover data collection and structure determination using serial femtosecond crystallography. For complete details on the use and execution of this protocol, please refer to Durdagi et al. (2021).
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Rosetta 2(DE3) Singles Competent Cells - Novagen, Novagen′s Rosetta 2 host strains are BL21 derivatives designed to enhance the expression of eukaryotic proteins that contain codons rarely used in E. coli.