6HAC

Crystal structure of [Fe]-hydrogenase (Hmd) holoenzyme from Methanococcus aeolicus (open form)

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	2016-09-26
Detector	ADSC QUANTUM 315r
Wavelength(s)	0.97980
Spacegroup name	I 41 2 2
Unit cell lengths	136.985, 136.985, 117.317
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	26.620 - 2.300
R-factor	0.172
Rwork	0.171
R-free	0.19200
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	4jjf
RMSD bond length	0.009
RMSD bond angle	1.110
Data reduction software	XDS
Data scaling software	SCALA (3.3.22)
Phasing software	PHASER
Refinement software	BUSTER (2.10.3)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	48.430	2.420
High resolution limit [Å]	2.300	2.300
Rmerge	0.155	1.023
Rmeas	0.161	1.061
Rpim	0.043	0.281
Number of reflections	25078	3607
<I/σ(I)>	17.9	2.9
Completeness [%]	100.0	100
Redundancy	13.7	14
CC(1/2)	0.999	0.602

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	4	281.15	The reconstituted [Fe]-hydrogenase holoenzyme from M. aeolicus was crystallized in the anaerobic tent with gas phase 95%N2/5%H2 using 96-well 2-drop MRC Crystallization Plates (Molecular Dimensions, Suffolk, UK). For the initial screening, 0.7-ul of 24 mg/ml reconstituted [Fe]-hydrogenase was mixed with 0.7 ul of reservoir solution of crystallization kits under yellow light and incubated under the dark condition. The best diffracting crystal was obtained in one month in the crystallization reservoir solution containing 20% w/v polyethylene glycol 3350, 100-mM tri-sodium citrate pH 4.0 and 200-mM tri-sodium citrate.