6YUA

CO-dehydrogenase coupled to the N-terminal domain of the Acetyl-CoA synthase from Clostridium autoethanogenum isolated after tryptic digestion.

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	SLS BEAMLINE X06DA
Synchrotron site	SLS
Beamline	X06DA
Temperature [K]	100
Detector technology	PIXEL
Collection date	2019-12-16
Detector	DECTRIS PILATUS 2M-F
Wavelength(s)	1.00000
Spacegroup name	P 41 21 2
Unit cell lengths	89.333, 89.333, 527.130
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	39.950 - 3.160
R-factor	0.2252
Rwork	0.224
R-free	0.25200
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	6ytt
Data reduction software	XDS
Data scaling software	STARANISO
Phasing software	MOLREP
Refinement software	PHENIX (1.17.1_3660)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	131.783	3.576
High resolution limit [Å]	3.140	3.140
Rmerge	0.260	1.492
Rmeas	0.268	1.524
Rpim	0.061	0.309
Number of reflections	17564	977
<I/σ(I)>	8.9	2.5
Completeness [%]	84.3	81.2
Redundancy	17.7	23.6
CC(1/2)	0.999	0.748

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	7.6	291.15	Crystallization was performed in an anoxic chamber (N2/H2, 95:5 %) with anaerobic solutions. Crystals were obtained by initial screening at 291.15 K using the sitting drop method on a 96-well MRC 2 Crystallization Plates in polystyrene (SWISSCI). The crystallization reservoir contained 90 uL of mother liquor, crystallization drop contained a mixture of 0.6 uL protein and 0.6 uL precipitant. The protein concentration was 28.5 mg/mL in 25 mM Tris pH 7.6, 10% (v/v) glycerol and 2 mM dithiothreitol. The best diffracting crystal of CODH bound to the ACS N-terminal domain was obtained by initial screening using the Shotgun (SG1) screen from Molecular dimensions. The crystallization reservoir contained 200 mM Magnesium chloride hexahydrate and 20% (w/v) polyethylene glycol 3350. The crystal was cryoprotected in the same crystallization solution supplemented with 25% ethylene glycol.