6V2N

Crystal structure of E. coli phosphoenolpyruvate carboxykinase mutant Lys254Ser

Replaces:  6CU4

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	CLSI BEAMLINE 08ID-1
Synchrotron site	CLSI
Beamline	08ID-1
Temperature [K]	105
Detector technology	CCD
Collection date	2014-07-10
Detector	RAYONIX MX300HE
Wavelength(s)	0.9793
Spacegroup name	P 1 21 1
Unit cell lengths	54.906, 76.647, 65.421
Unit cell angles	90.00, 96.38, 90.00

Refinement procedure

Resolution	44.290 - 1.650
R-factor	0.1465
Rwork	0.146
R-free	0.17430
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1oen
Data reduction software	XDS
Data scaling software	XDS
Phasing software	MOLREP
Refinement software	PHENIX (1.17.1_3660)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	44.450	1.709
High resolution limit [Å]	1.650	1.650
Rmerge	0.076	0.319
Rmeas	0.084	0.356
Rpim	0.036	0.158
Number of reflections	64733	6479
<I/σ(I)>	14.67	4.56
Completeness [%]	99.8	99.92
Redundancy	5.1	4.9
CC(1/2)	0.997	0.953

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	MICROBATCH		293	2 ul drop containing 2 mg/ml Lys254Ser E. coli PCK, 5 mM MnCl2, 5mM MgCl2, 2mM ATP, 2mM pyruvate, 1 mM EDTA, 200 mM ammonium acetate, 100 mM sodium acetate pH 4.8, 0.01 mM DTT and 10% PEG 4000, was added to 2 ul drop containing 0.2 M calcium chloride and 20% PEG. Rod like crystals formed after 7 days, were harvested, and soaked in cryoprotectant solution (30% glycerol, 1mM EDTA, 100 mM sodium acetate, 200 mM ammonium acetate and 12% PEG 4000) for 10 seconds and flash cooled in liquid nitrogen