To purify the RBD/ACE2 complex, human ACE2 and RBD were incubated together, and then the complex was purified on Superdex200 gel filtration chromatography. RBD/ACE2 crystals were grown in sitting drops at room temperature over wells containing 100 mM Tris (pH 8.5), 18-20% PEG 6000, and 100 mM NaCl. Crystals were soaked briefly in 100 mM Tris (pH 8.5), 30% PEG 6000, 100 mM NaCl, and 30% ethylene glycol before being flash-frozen in liquid nitrogen. X-ray diffraction data were collected at the Advanced Photon Source beamline 24-ID-E. The structure was determined by molecular replacement using the structure of human ACE2 complexed with SARS-CoV RBD as the search template (Protein Data Bank accession code 2AJF). Structure data and refinement statistics are shown in Extended Data Table.1 .
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SARS-CoV-2 RBD-ACE2 Complex Purification and Crystallization
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Ace2
Ethylene glycol
Frozen
Gel filtration chromatography
Human
Nacl
Nitrogen
Peg 6000
Sars cov
Tris
X ray diffraction
Corresponding Organization :
Other organizations : University of Minnesota
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Variable analysis
independent variables
- Incubation of human ACE2 and RBD together
- Purification of the RBD/ACE2 complex on Superdex200 gel filtration chromatography
- Growth of RBD/ACE2 crystals in sitting drops
- X-ray diffraction data collection at the Advanced Photon Source beamline 24-ID-E
- Determination of the structure by molecular replacement using the structure of human ACE2 complexed with SARS-CoV RBD as the search template
- 100 mM Tris (pH 8.5)
- 18-20% PEG 6000
- 100 mM NaCl
- 100 mM Tris (pH 8.5), 30% PEG 6000, 100 mM NaCl, and 30% ethylene glycol
- Structure of human ACE2 complexed with SARS-CoV RBD (Protein Data Bank accession code 2AJF)
- Not explicitly mentioned
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