9OET

Hydrox with succinate and vanadium(IV)-oxo

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ALS BEAMLINE 8.3.1
Synchrotron site	ALS
Beamline	8.3.1
Temperature [K]	90
Detector technology	PIXEL
Collection date	2022-08-30
Detector	DECTRIS PILATUS3 S 6M
Wavelength(s)	1.11583
Spacegroup name	C 2 2 21
Unit cell lengths	88.892, 139.970, 95.427
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	75.040 - 1.900
R-factor	0.2032
Rwork	0.201
R-free	0.24310
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.007
RMSD bond angle	0.893
Data reduction software	XDS (June 1, 2017)
Data scaling software	Aimless (0.7.9)
Phasing software	PHASER (2.8.3)
Refinement software	PHENIX (1.20.1_4487)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	95.390	1.968
High resolution limit [Å]	1.900	1.900
Rmerge	0.163	2.968
Rmeas	0.168	3.012
Rpim	0.040	0.512
Number of reflections	47209	4669
<I/σ(I)>	18.6	2.11
Completeness [%]	100.0	99.62
Redundancy	33.24	33.73
CC(1/2)	0.999	0.773

Crystallization Conditions

crystal 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)