6SDG

Crystal structure of the DNA binding domain of M. polymorpha Auxin Response Factor 2 (MpARF2) in complex with High Affinity DNA

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ALBA BEAMLINE XALOC
Synchrotron site	ALBA
Beamline	XALOC
Temperature [K]	100
Detector technology	PIXEL
Collection date	2017-11-24
Detector	DECTRIS PILATUS 6M
Wavelength(s)	0.97919
Spacegroup name	C 1 2 1
Unit cell lengths	162.768, 79.360, 79.767
Unit cell angles	90.00, 116.93, 90.00

Refinement procedure

Resolution	29.200 - 2.960
R-factor	0.2183
Rwork	0.215
R-free	0.27750
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	4ldx
RMSD bond length	0.013
RMSD bond angle	1.451
Data reduction software	XDS
Data scaling software	autoPROC
Phasing software	PHASER
Refinement software	PHENIX (1.16_3549)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	29.200	3.423
High resolution limit [Å]	2.960	2.960
Rmerge	0.089	1.641
Number of reflections	14035	1336
<I/σ(I)>	8.2	0.7
Completeness [%]	73.5	5.4
Redundancy	3.3	3.3

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	8	291	Required amount of DNA to 2:1 (protein:DNA) stoichiometry was diluted in [20 mM Tris (pH8.0), 500 mM NaCl] and protein was added to final concentration of 5 mg/mL. Crystals of MpARF2-DBD:DNA were obtained in 0.2 M Sodium formate, 20% w/v Polyethylene glycol 3,350 by the sitting-drop vapor-diffusion method at 18 deg. C, in drops of 1 uL + 1 uL (protein:reservoir). Cube-shaped crystals appeared in less than a week. Cryo-cooling in liquid nitrogen was carried out using a cryo-protecting solution containing reservoir solution supplemented with 10% glycerol