2VLF

Quinonoid intermediate of Citrobacter freundii tyrosine phenol-lyase formed with alanine

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	EMBL/DESY, HAMBURG BEAMLINE BW7B
Synchrotron site	EMBL/DESY, HAMBURG
Beamline	BW7B
Temperature [K]	120
Detector technology	IMAGE PLATE
Collection date	1995-10-07
Detector	MARRESEARCH
Spacegroup name	P 21 21 2
Unit cell lengths	132.846, 143.687, 59.618
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	20.000 - 1.890
R-factor	0.158
Rwork	0.157
R-free	0.19800
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	2ez2
RMSD bond length	0.016
RMSD bond angle	1.438
Data reduction software	DENZO
Data scaling software	SCALEPACK
Phasing software	AMoRE
Refinement software	REFMAC (5.3.0037)

Data quality characteristics

	Overall	Outer shell
Low resolution limit [Å]	20.000	1.930
High resolution limit [Å]	1.890	1.890
Rmerge	0.070	0.360
Number of reflections	89055
<I/σ(I)>	12.2
Completeness [%]	97.3	87.5
Redundancy	3.7	2.3

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, HANGING DROP	8	293	CRYSTALS OF THE C. FREUNDII TPL WERE GROWN AT 277 AND 293 K USING THE HANGING DROP VAPOR DIFFUSION METHOD. THE BEST CRYSTALS WERE OBTAINED BY MIXING 2 UL OF THE PROTEIN SOLUTION (18-20 MG/ML) CONTAINING 50MM K-PHOSPHATE PH 8.0, 0.5 MM PLP, 1MM DDT WITH AN EQUAL VOLUME OF THE RESERVOIR SOLUTION CONTAINING 50 MM TRIETHANOLAMINE BUFFER (PH 8.0), 0.5 MM PLP, 2 MM DDT, 0.4 M KCL, AND 35-38% (W/V) POLY(ETHYLENE GLYCOL) 5000 MONOMETHYL ETHER. QUINONOID INTERMEDIATE WITH ALANINE WAS PREPARED BY SOAKING THE TPL CRYSTALS FOR 5 MIN IN THE STABILIZATION SOLUTION CONTAINING 40% POLY(ETHYLENE GLYCOL) 5000 MONOMETHYL ETHER, 50 MM TRIETHANOLAMINE BUFFER (PH 8.0), 0.25 M KCL, 0.2 MM PLP, 0.5 MM DTT AND 10 MM L-ALA.