7KOA

Room Temperature Structure of SARS-CoV-2 Nsp10/16 Methyltransferase in a Complex with Cap-0 and SAM Determined by Pink-Beam Serial Crystallography

X-RAY DIFFRACTION

Serial Crystallography (SX)

Starting Model(s)

Initial Refinement Model(s)
Type	Source	Accession Code	Details
experimental model	PDB	6XKM

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	BATCH MODE	6.5	298	Protein: 4.0 mg/ml (NSP10/NSP16 1:1), 0.15 M NaCl, 0.01 M Tris-HCl, 2 mM SAM, 1 mM TCEP, 5% glycerol, pH 7.5.; Precipitation buffer: 0.1 M MES pH 6.5, 0.9 M NaF. Batch crystallization: 100 ul of protein mixed with 100 ul of precipitation buffer in 500 ul polypropylane tube. Crystals were soaked with m7GpppA (0.5 mM) for 10 minutes before data collection.

Crystal Properties
Matthews coefficient	Solvent content
4.46	72.5

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 170.86	α = 90
b = 170.86	β = 90
c = 52.797	γ = 120

Symmetry
Space Group	P 31 2 1

Diffraction

Diffraction Experiment
ID #	Crystal ID	Scattering Type	Data Collection Temperature	Detector	Detector Type	Details	Collection Date	Monochromator	Protocol
1	1	x-ray	298	CCD	RAYONIX MX340-HS		2020-09-18	L	LAUE

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	APS BEAMLINE 14-ID-B	1.02-1.18	APS	14-ID-B

Serial Crystallography

Sample delivery method
Diffraction ID	Description	Sample Delivery Method
1	Nylon Mesh	fixed target

Fixed Target
Diffraction ID	Description	Sample Holding	Support Base	Motion control	Details	Sample Solvent
1	ALEX mesh holder	nylon mesh	xyz stage	SmarAct Motors viaPMAC	start/stop raster over area	50 mM MES pH 6.5, 0.45 M NaF, 75 mM NaCl, 5 mM Tris-HCl, 1 mM SAM, 0.5 mM TCEP, 2.5 % glycerol, 0.5 mM m7GpppA

Data Reduction
Diffraction ID	Frames Indexed	Crystal Hits	Frames Indexed	Latices Merged
1	157	1817	157

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	R Merge I (Observed)	Net I Over Average Sigma (I)	Redundancy	Number Reflections (All)	Number Reflections (Observed)	Observed Criterion Sigma (F)	Observed Criterion Sigma (I)	B (Isotropic) From Wilson Plot
1	2.4	49.7	65.85	0.129	16.5	7.4		22795			20

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	R Merge I (Observed)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	2.4	2.51	15.63		0.105

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Starting model	Resolution (High)	Resolution (Low)	Number Reflections (Observed)	Number Reflections (R-Free)	Percent Reflections (Observed)	R-Factor (Observed)	R-Work	R-Free	R-Free Selection Details	Mean Isotropic B
X-RAY DIFFRACTION	MOLECULAR REPLACEMENT	THROUGHOUT	6XKM	2.4	29.59	21581	1198	65.79	0.2362	0.2339	0.2766	RANDOM	39.221

Temperature Factor Modeling
Anisotropic B[1][1]	Anisotropic B[1][2]	Anisotropic B[1][3]	Anisotropic B[2][2]	Anisotropic B[2][3]	Anisotropic B[3][3]
-0.01	-0.01		-0.01		0.04

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	21.064
r_dihedral_angle_3_deg	7.728
r_dihedral_angle_4_deg	6.979
r_dihedral_angle_1_deg	1.931
r_angle_refined_deg	1.105
r_angle_other_deg	0.264
r_gen_planes_refined	0.052
r_gen_planes_other	0.046
r_chiral_restr	0.043
r_bond_refined_d	0.003

RMS Deviations
Key	Refinement Restraint Deviation
r_dihedral_angle_2_deg	21.064
r_dihedral_angle_3_deg	7.728
r_dihedral_angle_4_deg	6.979
r_dihedral_angle_1_deg	1.931
r_angle_refined_deg	1.105
r_angle_other_deg	0.264
r_gen_planes_refined	0.052
r_gen_planes_other	0.046
r_chiral_restr	0.043
r_bond_refined_d	0.003
r_bond_other_d	0.001

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	3240
Nucleic Acid Atoms
Solvent Atoms	182
Heterogen Atoms	131

Software

Software
Software Name	Purpose
REFMAC	refinement
PDB_EXTRACT	data extraction
MOLREP	phasing
Precognition	data reduction
Epinorm	data scaling

RCSB PDB Core Operations are funded by the U.S. National Science Foundation (DBI-2321666), the US Department of Energy (DE-SC0019749), and the National Cancer Institute, National Institute of Allergy and Infectious Diseases, and National Institute of General Medical Sciences of the National Institutes of Health under grant R01GM157729.