6PT8

Crystal Structure of CobT from Methanocaldococcus jannaschii in complex with Adenine Alpha-Ribotide and Nicotinic Acid

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 19-ID
Synchrotron site	APS
Beamline	19-ID
Temperature [K]	100
Detector technology	PIXEL
Collection date	2018-08-15
Detector	DECTRIS PILATUS3 X 6M
Wavelength(s)	0.97934
Spacegroup name	P 1 21 1
Unit cell lengths	46.390, 138.640, 51.150
Unit cell angles	90.00, 108.32, 90.00

Refinement procedure

Resolution	41.973 - 1.400
R-factor	0.1542
Rwork	0.153
R-free	0.17480
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	3l0z
Data reduction software	XDS
Data scaling software	XSCALE
Phasing software	PHASER
Refinement software	PHENIX (1.15.2_3472)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	41.973	1.450
High resolution limit [Å]	1.400	1.400
Rmerge	0.096	0.667
Number of reflections	118689	11857
<I/σ(I)>	12.18	2.92
Completeness [%]	99.0	98.98
Redundancy	6.8	6.8

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP		298	Crystals formed by mixing 1:1 ratio of 13.3 mg/mL protein solution with well solution containing 100 mM 4-(2-Hydroxyethyl)-1-piperazinepropanesulfonic acid (HEPPS) pH 8.5 and 22% (w/v) PEG 8K. 1 mM adenine and 50 mM Sodium/Potassium Phosphate pH 7.0 were added to protein prior to mixing with well solution. Substrates were captured in lattice prior to freezing by soaking crystals in 20% (w/v) ethylene glycol, 100 mM HEPPS pH 8.5, 22% (w/v) PEG 8K, 25 mM sodium/potassium phosophate pH 7.0, 1 mM nicotinic acid mononucleotide, and 1 mM adenine for 30 minutes. Crystals were then soaked in the same solution, but with 30% PEG 8K instead of 20% ethylene glycol and 20% PEG 8K