7ROO

Crystal structure of Friedel-Crafts alkylating enzyme CylK from Cylindospermum licheniforme with bromide

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	APS BEAMLINE 21-ID-G
Synchrotron site	APS
Beamline	21-ID-G
Temperature [K]	80
Detector technology	CCD
Collection date	2020-07-09
Detector	MARMOSAIC 300 mm CCD
Wavelength(s)	0.97857
Spacegroup name	C 2 2 21
Unit cell lengths	93.470, 138.656, 99.511
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	46.780 - 1.520
R-factor	0.1708
Rwork	0.169
R-free	0.19780
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	7ron
RMSD bond length	0.013
RMSD bond angle	1.823
Data reduction software	HKL-2000
Data scaling software	SCALEPACK
Phasing software	REFMAC
Refinement software	REFMAC (5.8.0258)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	50.000	1.550
High resolution limit [Å]	1.520	1.520
Number of reflections	98866	4856
<I/σ(I)>	23.3	2.14
Completeness [%]	99.7
Redundancy	7	5.5
CC(1/2)	0.990	0.843

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	7	298	Prior to crystallization trials, purified CylK protein in storage buffer (20 mM HEPES pH 7.8, 50 mM NaCl, 10 mM MgCl2, 10 mM CaCl2, 10% glycerol) was combined with 5-undecylbenzene-1,3-diol (UDR), a substrate analog dissolved in protein storage buffer containing 22% DMSO. The final protein solution contained 10 mg/mL CylK and 1.66 mM UDR. Crystals of undecylresorcinol-bound CylK were obtained by using the hanging drop vapor diffusion method in 2 uL drops mixed in 1:1 ratio with a precipitant solution of 1.8 M sodium malonate, pH 7.0. Crystals formed within about 1 week at room temperature. CylK crystals were soaked in 2 M sodium malonate with 500 mM NaBr for up to 1 min