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Protocol for structure determination of SARS-CoV-2 main protease at near-physiological-temperature by serial femtosecond crystallography.
Ertem, Fatma Betul; Guven, Omur; Buyukdag, Cengizhan; Gocenler, Oktay; Ayan, Esra; Yuksel, Busra; Gul, Mehmet; Usta, Gozde; Cakilkaya, Baris; Johnson, J Austin; Dao, E Han; Su, Zhen; Poitevin, Frederic; Yoon, Chun Hong; Kupitz, Christopher; Hayes, Brandon; Liang, Mengning; Hunter, Mark S; Batyuk, Alexander; Sierra, Raymond G; Ketawala, Gihan; Botha, Sabine; Dag, Çagdas; DeMirci, Hasan.
  • Ertem FB; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Guven O; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Buyukdag C; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Gocenler O; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Ayan E; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Yuksel B; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Gul M; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Usta G; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Cakilkaya B; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Johnson JA; Department of Molecular Biology and Genetics, Koc University, Istanbul, Turkey.
  • Dao EH; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Su Z; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Poitevin F; Department of Applied Physics, Stanford University, Stanford, CA, USA.
  • Yoon CH; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Kupitz C; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Hayes B; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Liang M; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Hunter MS; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Batyuk A; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Sierra RG; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Ketawala G; Linac Coherent Light Source, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, 94025 CA, USA.
  • Botha S; Department of Physics, Arizona State University, Tempe, AZ 85287-1504, USA.
  • Dag Ç; Biodesign Center for Applied Structural Discovery, Arizona State University, Tempe, AZ 85287-5001, USA.
  • DeMirci H; Department of Physics, Arizona State University, Tempe, AZ 85287-1504, USA.
STAR Protoc ; 3(1): 101158, 2022 03 18.
Article in English | MEDLINE | ID: covidwho-1650422
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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Full text: Available Collection: International databases Database: MEDLINE Main subject: Models, Molecular / Coronavirus 3C Proteases / SARS-CoV-2 Limits: Humans Language: English Journal: STAR Protoc Year: 2022 Document Type: Article Affiliation country: J.xpro.2022.101158

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Models, Molecular / Coronavirus 3C Proteases / SARS-CoV-2 Limits: Humans Language: English Journal: STAR Protoc Year: 2022 Document Type: Article Affiliation country: J.xpro.2022.101158