7Z59

SARS-CoV-2 main protease (Mpro) covalently modified with a penicillin derivative

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	DIAMOND BEAMLINE I03
Synchrotron site	Diamond
Beamline	I03
Temperature [K]	100
Detector technology	PIXEL
Collection date	2021-12-11
Detector	DECTRIS EIGER X 16M
Wavelength(s)	0.97625
Spacegroup name	C 1 2 1
Unit cell lengths	113.341, 53.808, 44.703
Unit cell angles	90.00, 100.65, 90.00

Refinement procedure

Resolution	55.695 - 2.000
Rwork	0.222
R-free	0.25160
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	6yb7
RMSD bond length	0.008
RMSD bond angle	1.481
Data reduction software	xia2
Data scaling software	Aimless
Phasing software	MOLREP
Refinement software	REFMAC (5.8.0267)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	55.695	55.690	2.050
High resolution limit [Å]	2.000	8.940	2.000
Rmerge	0.188	0.047	4.066
Rmeas	0.208	0.051	4.490
Rpim	0.088	0.021	1.893
Number of reflections	18025	2192	14239
<I/σ(I)>	7.9
Completeness [%]	100.0	100	100
Redundancy	10.7	10.1	10.8
CC(1/2)	0.997	0.998	0.211

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	6	293	SARS-CoV-2 Mpro was thawed and diluted to 6 mg/mL (using 20 mM HEPES, pH 7.5, 50 mM NaCl). Beta-Lactam 20e was added to the protein solution to a final concentration of 10 mM; the mixture was incubated for 2 h at ambient temperature prior to dispensing plates. The drop composition was 0.15 uL protein ligand solution, 0.3 uL 11 percent v/v PEG 4000, 0.1 M MES, pH 6.5, and 0.05 ,microL Mpro crystal seed stock. The Mpro crystal seed stock was prepared by crushing Mpro crystals with a pipette tip, suspending them in 30% PEG 4000, 5%v/v DMSO, 0.1 M MES pH 6.5, and vortexing for 60 s with approximately 10 glass beads (1.0 mm diameter, BioSpec products). As reservoir solution was used: 11%v/v PEG 4K, 5%v/v DMSO, 0.1 M MES, pH 6.5. Crystals were grown using the sitting drop vapor diffusion method at 20 degree C and appeared within 24 h, reaching full size within 36 h. Crystals were looped after one week.