9TEI

Fosfomycin-resistance protein from Klebsiella pneumoniae (FosAKP) crystal structure by beam sweeping serial crystallography

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	FREE ELECTRON LASER
Source details	EUROPEAN XFEL BEAMLINE SPB/SFX
Synchrotron site	European XFEL
Beamline	SPB/SFX
Temperature [K]	298
Detector technology	PIXEL
Collection date	2024-09-23
Detector	PSI JUNGFRAU 4M
Wavelength(s)	1.36245
Spacegroup name	P 21 21 2
Unit cell lengths	71.170, 91.410, 45.290
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	35.590 - 2.000
R-factor	0.1811
Rwork	0.176
R-free	0.22330
Structure solution method	MOLECULAR REPLACEMENT
RMSD bond length	0.007
RMSD bond angle	0.996
Data reduction software	CrystFEL (0.11.1)
Data scaling software	CrystFEL (0.11.1)
Phasing software	PHASER (2.8.3)
Refinement software	PHENIX (1.21.2_5419)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	35.590	2.050
High resolution limit [Å]	2.000	2.000
Number of reflections	20629	1455
<I/σ(I)>	7.91	1.57
Completeness [%]	100.0	99.93
Redundancy	476.9	82.1
CC(1/2)	0.950	0.560

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP		296	25 mg/mL FosAKP in 10 mM Hepes, pH 7.5, 75 mM NaCl was supplemented with 6 mM MnCl2 and mixed with an equal volume of 16% (w/v) PEG3350, 0.25 M MgCl2, 0.2 M KBr, 0.1 M BisTris, pH 5.5 and 1/10 volume of seed stock in 26% (w/v) PEG3350, 0.25 M MgCl2, 0.2 M KBr, 0.1 M BisTris, pH 5.5. From this, approximately 66 uL were added per window of the fixed-target chip. The sample holder was then inserted into a 3D-printed crystal growth chamber with 3 mL of precipitant solution in the bottom for vapor-diffusion crystallization and incubated at 20 degrees C. For data collection excess precipitant was removed by blotting and the sample holders were equipped with a protective cover to prevent them from drying-out.