5LEK

The Transcriptional Regulator PrfA-G145S mutant from Listeria Monocytogenes in complex with a 30-bp operator PrfA-box motif

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	BESSY BEAMLINE 14.1
Synchrotron site	BESSY
Beamline	14.1
Temperature [K]	100
Detector technology	PIXEL
Collection date	2016-01-22
Detector	DECTRIS PILATUS 6M-F
Wavelength(s)	0.8729
Spacegroup name	P 43 21 2
Unit cell lengths	79.081, 79.081, 265.978
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	47.297 - 2.800
R-factor	0.233
Rwork	0.231
R-free	0.26660
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	2bgc
RMSD bond length	0.004
RMSD bond angle	0.761
Data reduction software	XDS
Data scaling software	Aimless
Phasing software	PHASER
Refinement software	PHENIX ((1.10.1_2155: ???))

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	47.300	2.950
High resolution limit [Å]	2.800	2.800
Rmerge	0.091	1.281
Number of reflections	21616
<I/σ(I)>	14	1.7
Completeness [%]	99.4	99.2
Redundancy	7	6.9

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	5.5	291	Protein and duplex DNA were incubated together at a ratio of 1:1.3 (PrfA dimer:hly DNA) at final concentrations of 50 microM and 70 microM respectively in 20 mM Tris-HCl pH 8.0, 150 mM NaCl, 1 mM DTT for 60 min at room temperature, before being used for crystal setups. Crystals were obtained after 24 h by mixing 4 microL protein-DNA solution with 2 microL reservoir solution consisting of 8% PEG 8000, 100 mM sodium acetate pH 4.6, 100 mM magnesium acetate, 20% glycerol. Prior to vitrification the soaking of PrfAG145S-DNA crystals were soaked in a reservoir solution containing 30% glycerol for 24 h