Experiment: 9OET

X-RAY DIFFRACTION

Starting Model(s)

Initial Refinement Model(s)
Type	Source	Accession Code	Details
experimental model	PDB	7JSD

Crystallization

Crystalization Experiments
ID	Method	pH	Temperature	Details
1	VAPOR DIFFUSION, HANGING DROP		291	Hydrox crystals were obtained by the hanging drop vapor diffusion method by combining equal volumes of protein solution (Hydrox (5 mg/ml), lysine (50 mM, pH 7), sodium succinate (20 mM, pH 7), and vanadium(IV) oxide sulfate hydrate (1 mM)) and reservoir solution (HEPES (100 mM), 16% (w/v) PEG 6000, magnesium chloride hexahydrate (200 mM), pH 9). Crystals grew in three days and were transferred to an Eppendorf tube containing 250 uL of reservoir solution and vortexed for 30 seconds with 10 x 1 um diameter zirconia/silica beads to produce a micro-seed solution. The seed solution was stored at -80C for future use. Subsequent crystals were prepared by micro-seeding equal volumes of protein solution (Hydrox (5 mg/ml), lysine (50 mM, pH 7), sodium succinate (20 mM, pH 7), and vanadium(IV) oxide sulfate hydrate (1 mM)) and reservoir solution (HEPES (100 mM), 16% (w/v) PEG 6000, magnesium chloride hexahydrate (200 mM), pH 9)

Crystal Properties
Matthews coefficient	Solvent content
2.49	50.63

Crystal Data

Unit Cell
Length ( Å )	Angle ( ˚ )
a = 88.892	α = 90
b = 139.97	β = 90
c = 95.427	γ = 90

Symmetry
Space Group	C 2 2 21

Diffraction

Diffraction Experiment
ID #	Crystal ID	Scattering Type	Data Collection Temperature	Detector	Detector Type	Details	Collection Date	Monochromator	Protocol
1	1	x-ray	90	PIXEL	DECTRIS PILATUS3 S 6M		2022-08-30	M	SINGLE WAVELENGTH

Radiation Source
ID #	Source	Type	Wavelength List	Synchrotron Site	Beamline
1	SYNCHROTRON	ALS BEAMLINE 8.3.1	1.11583	ALS	8.3.1

Data Collection

Overall
ID #	Resolution (High)	Resolution (Low)	Percent Possible (Observed)	R Merge I (Observed)	Rrim I (All)	Rpim I (All)	CC (Half)	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	1.9	95.39	100	0.163	0.168	0.04	0.999	18.6	33.24		47209

Highest Resolution Shell
ID #	Resolution (High)	Resolution (Low)	Percent Possible (All)	Percent Possible (Observed)	R Merge I (Observed)	Rrim I (All)	Rpim I (All)	CC (Half)	Mean I Over Sigma (Observed)	Redundancy	Number Unique Reflections (All)
1	1.9	1.968	99.62		2.968	3.012	0.5123	0.773	2.11	33.73

Refinement

Statistics
Diffraction ID	Structure Solution Method	Cross Validation method	Resolution (High)	Resolution (Low)	Cut-off Sigma (F)	Number Reflections (Observed)	Number Reflections (R-Free)	Percent Reflections (Observed)	R-Factor (Observed)	R-Work (Depositor)	R-Work (DCC)	R-Free (Depositor)	R-Free (DCC)	Mean Isotropic B
X-RAY DIFFRACTION	MOLECULAR REPLACEMENT	FREE R-VALUE	1.9	75.04	1.34	47179	2424	99.92	0.2032	0.201	0.2013	0.2431	0.2433

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]

RMS Deviations
Key	Refinement Restraint Deviation
f_dihedral_angle_d	14.321
f_angle_d	0.893
f_chiral_restr	0.056
f_plane_restr	0.015
f_bond_d	0.007

Non-Hydrogen Atoms Used in Refinement
Non-Hydrogen Atoms	Number
Protein Atoms	3367
Nucleic Acid Atoms
Solvent Atoms	425
Heterogen Atoms	12

Software

Software
Software Name	Purpose
PHENIX	refinement
PDB_EXTRACT	data extraction
XDS	data reduction
Aimless	data scaling
PHASER	phasing

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.