6COM

2.3A crystal structure of E. coli phosphoenolpyruvate carboxykinase mutant Asp269Asn

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	CLSI BEAMLINE 08B1-1
Synchrotron site	CLSI
Beamline	08B1-1
Temperature [K]	105
Detector technology	CCD
Collection date	2014-02-28
Detector	RAYONIX MX300HE
Wavelength(s)	0.98
Spacegroup name	C 1 2 1
Unit cell lengths	124.595, 94.175, 46.457
Unit cell angles	90.00, 94.77, 90.00

Refinement procedure

Resolution	47.087 - 2.300
R-factor	0.1535
Rwork	0.148
R-free	0.20920
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1ayl
RMSD bond length	0.008
RMSD bond angle	0.963
Data reduction software	HKL-2000
Data scaling software	d*TREK
Phasing software	PHASER
Refinement software	PHENIX (dev_2398)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	50.000	2.190
High resolution limit [Å]	2.110	2.110
Rmerge	0.250
Number of reflections	77150
<I/σ(I)>	5.8
Completeness [%]	99.8
Redundancy	3.5

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	MICROBATCH	8.2	293	A 2ul drop with 2 mg/ml protein, 5 mM Calcium chloride, 5mM Magnesium chloride, 2 mM ATP, 2mM Pyruvate, 1 mM EDTA, 200 mM Ammonium acetate, 100 mM Sodium acetate, 0.01 mM DTT, 30% PEG 4000 was added to 2 ul drop containing 2 M sodium aceate, 0.1 M Tris pH 8.2 30% PEG 4000. After a week a rod like crystal was removed and soaked in a solution with 30% glycerol 1mM EDTA, 100 mM sodium acetate 200mM ammonium acetate and 12% PEG 4000. The crystal was put into a loop and flash cooled in liquid notrogen