7ZQ0

Room temperature SSX structure of GH11 xylanase from Nectria haematococca (1000 frames)

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	PETRA III, DESY BEAMLINE P11
Synchrotron site	PETRA III, DESY
Beamline	P11
Temperature [K]	295
Detector technology	PIXEL
Collection date	2019-04-16
Detector	DECTRIS PILATUS 6M
Wavelength(s)	1.033
Spacegroup name	C 1 2 1
Unit cell lengths	80.550, 38.850, 53.570
Unit cell angles	90.00, 91.00, 90.00

Refinement procedure

Resolution	16.920 - 1.900
R-factor	0.2284
Rwork	0.226
R-free	0.26170
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	6y0h
RMSD bond length	0.004
RMSD bond angle	0.626
Data reduction software	CrystFEL
Data scaling software	CrystFEL
Phasing software	PHENIX
Refinement software	PHENIX (1.17.1_3660)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	16.920	1.930
High resolution limit [Å]	1.900	1.900
Number of reflections	13235	1325
<I/σ(I)>	1.91	0.68
Completeness [%]	99.7	99.33
Redundancy	20.98	13.2
CC(1/2)	0.751	0.185

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	BATCH MODE		293	For crystallization, the original conditions were modified to obtain microcrystals. Initial crystals obtained from hanging drops under the precipitant condition: 1 M ammonium sulfate, 100 mM sodium citrate pH 5.5 were crushed under a stereomicroscope, using a crystal crusher tool (Hampton research). The reservoir solution was pipetted to the drop and the seed stock was collected by washing the drop with reservoir solution. The seed stock was transferred to a seed bead tube (Molecular Dimensions Ltd., UK), vortexed three times for 30 s each, with an interval of 30 s between each vortex, to get the final seed stock. Protein solution 15 mg/mL, precipitant solution, and seed stock were mixed with a ratio of 1:1:0.5. The mixture was vortexed for 30 s in ten-minute intervals. After 30 min, the microcrystals were centrifuged at 200 rpm and the supernatant was replaced with a precipitant solution. Applying the same protocol, microcrystals were obtained under two precipitant conditions i.e. Precipitant 1: 1M (NH4)2SO4, 100 mM sodium citrate pH 5.5, and Precipitant 2: 200 mM (NH4)2SO4, 100 mM sodium citrate pH 5.5 and 20% PEG 6000. Microcrystals obtained under both precipitant conditions were tested for diffraction data collection