5TJO

Crystal structure of GTB + B trisaccharide (mercury derivative)

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	ROTATING ANODE
Source details	RIGAKU MICROMAX-002
Temperature [K]	113
Detector technology	IMAGE PLATE
Collection date	2009-04-16
Detector	RIGAKU RAXIS IV++
Wavelength(s)	1.5418
Spacegroup name	C 2 2 21
Unit cell lengths	52.710, 150.650, 79.180
Unit cell angles	90.00, 90.00, 90.00

Refinement procedure

Resolution	19.920 - 1.570
R-factor	0.1747
Rwork	0.174
R-free	0.19580
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	1lz7
RMSD bond length	0.010
RMSD bond angle	1.500
Data reduction software	d*TREK
Data scaling software	d*TREK (8.0SSI)
Phasing software	PHASER
Refinement software	REFMAC (5.8.0155)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	19.920	19.920	1.630
High resolution limit [Å]	1.570	3.380	1.570
Rmerge	0.040	0.029	0.317
Rmeas	0.040
Total number of observations	196681
Number of reflections	43816
<I/σ(I)>	17.3	41.1	3.7
Completeness [%]	98.6	94.7	98.1
Redundancy	4.46	4.89	3.7

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION		277	The first crystals of GTA/GTB were grown as mercury derivatives (Patenaude, S.I., et al. 2002): 10 ul drops with 6-8 mg/ml protein, 70 mM N-(2-acetamido)-2-iminodiacetic acid (ADA) pH 7.5, 50 mM, sodium acetate pH 4.6, 40 mM NaCl, 5-8 mM MnCl2, 2.5% (v/v) 2-methyl-2,4-pentanediol (MPD), 5%(v/v) glycerol, 2%(w/v) PEG 4000, and 0.3-0.5 mM 3-chloromercuri-2-methoxypropylurea suspended over 1 ml of a resevoir solution: 50 mM ADA pH 7.5, 10 mM MnCl2, 100 mM ammonium sulfate, 5%(v/v) MPD, 10%(v/v) glycerol, and 8-10%(w/v) PEG 4000. Crystals were washed with artificial mother liquor ML-1 consisting of 10% polyethylene glycol (PEG) 4000, 30 mM N-(2-acetamido)-2-iminodiacetic acid (ADA) buffer pH 7.5, 30 mM sodium acetate buffer pH 4.6, 100 mM ammonium sulfate, 10 mM MnCl2 and 30% glycerol as the cryoprotectant. Crystals of GTA/GTB in complex with the A and B trisaccharides were obtained by soaking them in mother liquor ML-1 with 30% glycerol and 45-50 mM substrates for 2-5 days at 18 degrees Celsius