5LRS

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

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ESRF BEAMLINE ID29
Synchrotron site	ESRF
Beamline	ID29
Temperature [K]	100
Detector technology	PIXEL
Collection date	2016-07-06
Detector	DECTRIS PILATUS3 6M
Wavelength(s)	1.073
Spacegroup name	P 43 21 2
Unit cell lengths	78.961, 78.961, 265.224
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	54.636 - 2.900
R-factor	0.2495
Rwork	0.248
R-free	0.27920
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	2bgc
RMSD bond length	0.003
RMSD bond angle	0.530
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]	58.900	3.080
High resolution limit [Å]	2.900	2.900
Rmerge	0.176	1.218
Number of reflections	19427
<I/σ(I)>	16	3.1
Completeness [%]	99.5	99.8
Redundancy	25.6	26.2

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	4.6	291	Prior to the crystallization setup GSH and DTT were added to the protein solution to final concentrations of 5 mM and 1 mM, respectively. 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 PrfAWT-DNA crystals were soaked in a reservoir solution containing 30% glycerol and 100 mM GSH for 24 h.