9R51

Dimeric state of the F420-reducing hydrogenase from Methanothermococcus thermolithotrophicus in crystalline form 1

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ESRF BEAMLINE BM30A
Synchrotron site	ESRF
Beamline	BM30A
Temperature [K]	100
Detector technology	CCD
Collection date	2016-09-23
Detector	ADSC QUANTUM 315r
Wavelength(s)	1.3871
Spacegroup name	P 32 2 1
Unit cell lengths	192.710, 192.710, 386.212
Unit cell angles	90.00, 90.00, 120.00

Refinement procedure

Resolution	31.550 - 2.300
R-factor	0.1812
Rwork	0.180
R-free	0.20740
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.007
RMSD bond angle	1.005
Data reduction software	XDS
Data scaling software	SCALA
Phasing software	PHASER
Refinement software	PHENIX ((1.20.1_4487: ???))

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	48.860	2.420
High resolution limit [Å]	2.300	2.300
Rmerge	0.091	0.597
Rmeas	0.100	0.673
Rpim	0.042	0.304
Number of reflections	365266	52491
<I/σ(I)>	13.6	2.8
Completeness [%]	99.8	99.1
Redundancy	5.7	4.7
CC(1/2)	0.998	0.565

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	7.5	293.15	The enzyme used for crystallization was at a concentration of 16 mg/ml in 25 mM Tris/HCl pH 7.6, 10 % (v/v) glycerol, 2 mM FAD and 2 mM dithiothreitol. Crystals were obtained anaerobically in a Coy tent filled with a N2/H2 (97:3%) atmosphere by initial screening at 20 degrees Celsius using the sitting drop method on 96-well MCR two-drop crystallization plates in polystyrene (SWISSCI). The reservoir contained 90 ul of crystallization solution. The crystallization solution contained 45 % (w/v) Pentaerythritol ethoxylate (3/4 EO/OH) 270, 100 mM HEPES pH 7.5, and 200 mM (NH4)2SO4. Drops of 0.7 ul protein and 0.7 ul crystallization solution were spotted.