8OJ1

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	2018-12-11
Detector	DECTRIS PILATUS 6M
Wavelength(s)	0.97926
Spacegroup name	I 21 21 21
Unit cell lengths	79.911, 80.933, 146.623
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	70.860 - 2.570
R-factor	0.2433
Rwork	0.242
R-free	0.25920
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.003
RMSD bond angle	0.539
Phasing software	PHASER
Refinement software	PHENIX (1.19.2_4158)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	70.860	2.610
High resolution limit [Å]	2.566	2.566
Rmerge	0.063	1.449
Rmeas	0.066
Number of reflections	15573	42
<I/σ(I)>	20	1.5
Completeness [%]	99.8
Redundancy	9.7
CC(1/2)	0.999	0.715

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP		293	Required amount of DNA to 2:1 (protein:DNA) stoichiometry was diluted in [20 mM Tris (pH8.0), 500 mM NaCl, 20 mM DTT] and protein was added to final concentration of 5 mg/mL. Crystals of MpARF2-DBD:DNA were obtained in 100 mM Tris pH 7-9, 18-33% PEG2KMME, 18.5% Glycerol by the sitting-drop vapor-diffusion method at 18 deg. C, in drops of 1 uL + 1 uL (protein:reservoir). 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.