6YKB

[Fe]-hydrogenase from Methanolacinia paynteri in complex with GMP at 1.55-A resolution

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)	0.97972
Spacegroup name	P 1 21 1
Unit cell lengths	67.805, 57.058, 95.125
Unit cell angles	90.00, 91.43, 90.00

Refinement procedure

Resolution	22.690 - 1.550
R-factor	0.185
Rwork	0.184
R-free	0.20900
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	4jjf
RMSD bond length	0.010
RMSD bond angle	1.100
Data reduction software	XDS
Data scaling software	SCALA (3.3.22)
Phasing software	MOLREP
Refinement software	BUSTER (2.10.3)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	95.095	39.915	1.630
High resolution limit [Å]	1.550	4.900	1.550
Rmerge		0.032	0.353
Rmeas	0.080	0.038	0.413
Rpim	0.041	0.020	0.213
Number of reflections	104812	3332	15285
<I/σ(I)>	12.6	18.8	2.2
Completeness [%]	99.3	96.1	99.7
Redundancy	3.7	3.6	3.7
CC(1/2)	0.997		0.653

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	7	283.15	Fe-hydrogenase holoenzyme from Methanolacinia paynteri was crystallized under 95%N2/5%H2 at 8 degree Celsius using a 96-well two-drop MRC crystallization plates. 0.7 ul of 25 mg/ml reconstituted enzyme with an Fe(II)-complex mimicking the FeGP cofactor and GMP (2mM final) was mixed with 0.7-ul reservoir solution under yellow light and incubated under dark conditions. The best crystal appeared within one month in 20% w/v polyethylene glycol 3350 and 200-mM magnesium formate reservoir solution (JBScreen Wizard 3&4 HTS, Jena Bioscience). The cryo protection was performed by adding 30% glycerol final to the crystallization solution.