9QV2

1-Phosphofructokinase mutant K95H from E. coli with bound fructose 6-phosphate and ADP

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ESRF BEAMLINE ID23-2
Synchrotron site	ESRF
Beamline	ID23-2
Temperature [K]	100
Detector technology	PIXEL
Collection date	2021-07-09
Detector	DECTRIS PILATUS3 2M
Wavelength(s)	0.873128
Spacegroup name	I 2 2 2
Unit cell lengths	63.560, 99.080, 140.620
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	29.380 - 1.850
R-factor	0.1604
Rwork	0.159
R-free	0.18930
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.019
RMSD bond angle	1.320
Data reduction software	XDS
Data scaling software	XSCALE
Phasing software	PHENIX
Refinement software	PHENIX ((1.21.2_5419: ???))

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	29.380	29.380	1.900
High resolution limit [Å]	1.850	8.270	1.850
Rmerge	0.083	0.031	0.641
Rmeas	0.090	0.034	0.688
Number of reflections	38279	460	2809
<I/σ(I)>	17.09
Completeness [%]	99.9
Redundancy	7.59
CC(1/2)	0.999	0.999	0.903

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP		289	The FruK variant K95H (3.4 mg/mL) in 20 mM MOPS/KOH buffer (pH 7.4) containing 75 mM KCl and 1 M L-proline was mixed in a 1:1 ratio with condition containing 1 M ammonium sulfate, 75 mM TRIS/HCl (pH 8.5), 150 mM lithium sulfate, 8 mM fructose-6-phosphate, 4 mM ADP, and 10 mM magnesium chloride. The final drop volume was 0.8 microliters. Before plunge freezing in liquid nitrogen, the crystals were soaked with additional 25 mM fructose-6-phosphate, 12.5 mM magnesium chloride for about 1 minute, before glycerol was added as a cryo-protectant to a final concentration of 30% (v/v).