5TJN
Crystal structure of GTB + B trisaccharide (native)
X-RAY DIFFRACTION
Starting Model(s)
Initial Refinement Model(s) | |||
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Type | Source | Accession Code | Details |
experimental model | PDB | 1LZ7 |
Crystallization
Crystalization Experiments | ||||
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ID | Method | pH | Temperature | Details |
1 | VAPOR DIFFUSION | 277 | Native crystals of GTA/GTB lacking any heavy metals were grown at 4 degrees Celsius from much higher concentrations of protein (30-40 mg/mL for GTB and 16-20 mg/mL for GTA) along with 1% PEG 4000, 4.5-5% 2-methyl-2,4-pentanediol (MPD), 100 mM ammonium sulfate, 70 mM sodium chloride, 50 mM ADA buffer pH 7.5, 30 mM sodium acetate buffer pH 4.6 and 5 mM MnCl2 for GTB crystallization and 5-8 mM CoCl2 for GTA crystallization. 10-15 microlitre drops were placed against a reservoir containing 3.7% PEG 4000, 7% MPD, 0.3 M ammonium sulfate, 0.25 M sodium chloride, 0.2 M ADA buffer and 0.1 M sodium acetate. The crystals were usually grown for 5-10 days. Before making complexes, crystals of GTA/GTB were washed with modified mother liquor ML-2 consisting of 3.5% PEG 4000, 50 mM ammonium sulfate, 40 mM sodium chloride, 35 mM ADA buffer and 15% MPD or glycerol. Crystals of native GTA/GTB in complex with the respective A or B trisaccharide were obtained by soaking them in mother liquor ML-2 with 15% glycerol or MPD and 45-50 mM of each substrate for 2-5 days at 4 degrees Celsius. Before freezing the crystals for data collection, the concentration of the cryoprotectant was made 30% glycerol or 20% MPD respectively |
Crystal Properties | |
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Matthews coefficient | Solvent content |
2.26 | 45.55 |
Crystal Data
Unit Cell | |
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Length ( Å ) | Angle ( ˚ ) |
a = 52.67 | α = 90 |
b = 150.03 | β = 90 |
c = 79.25 | γ = 90 |
Symmetry | |
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Space Group | C 2 2 21 |
Diffraction
Diffraction Experiment | ||||||||||||||
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ID # | Crystal ID | Scattering Type | Data Collection Temperature | Detector | Detector Type | Details | Collection Date | Monochromator | Protocol | |||||
1 | 1 | x-ray | 277 | IMAGE PLATE | RIGAKU RAXIS IV++ | 2009-04-16 | M | SINGLE WAVELENGTH |
Radiation Source | |||||
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ID # | Source | Type | Wavelength List | Synchrotron Site | Beamline |
1 | ROTATING ANODE | RIGAKU MICROMAX-002 | 1.5418 |
Data Collection
Overall | |||||||||||||||||||
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ID # | Resolution (High) | Resolution (Low) | Percent Possible (Observed) | R Merge I (Observed) | Net I Over Average Sigma (I) | Redundancy | Number Reflections (All) | Number Reflections (Observed) | Observed Criterion Sigma (F) | Observed Criterion Sigma (I) | B (Isotropic) From Wilson Plot | ||||||||
1 | 1.47 | 19.85 | 99.2 | 0.043 | 15.1 | 4.01 | 53360 |
Highest Resolution Shell | |||||||||||||||||||
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ID # | Resolution (High) | Resolution (Low) | Percent Possible (All) | Percent Possible (Observed) | R Merge I (Observed) | Mean I Over Sigma (Observed) | Redundancy | Number Unique Reflections (All) | |||||||||||
1 | 1.47 | 1.52 | 97.7 | 0.348 | 3.1 | 3.31 |
Refinement
Statistics | |||||||||||||||||||
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Diffraction ID | Structure Solution Method | Cross Validation method | Starting model | Resolution (High) | Resolution (Low) | Number Reflections (Observed) | Number Reflections (R-Free) | Percent Reflections (Observed) | R-Factor (Observed) | R-Work (Depositor) | R-Work (DCC) | R-Free (Depositor) | R-Free (DCC) | R-Free Selection Details | Mean Isotropic B | ||||
X-RAY DIFFRACTION | MOLECULAR REPLACEMENT | THROUGHOUT | 1LZ7 | 1.47 | 19.85 | 50642 | 2706 | 99.19 | 0.1843 | 0.1832 | 0.18 | 0.2044 | 0.2 | RANDOM | 21.136 |
Temperature Factor Modeling | ||||||
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Anisotropic B[1][1] | Anisotropic B[2][2] | Anisotropic B[3][3] | ||||
-0.58 | 0.32 | 0.26 |
RMS Deviations | |
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Key | Refinement Restraint Deviation |
r_dihedral_angle_2_deg | 32.067 |
r_dihedral_angle_4_deg | 15.915 |
r_dihedral_angle_3_deg | 10.952 |
r_dihedral_angle_1_deg | 6.39 |
r_mcangle_it | 1.844 |
r_angle_refined_deg | 1.505 |
r_mcbond_it | 1.23 |
r_mcbond_other | 1.225 |
r_angle_other_deg | 1.129 |
r_chiral_restr | 0.091 |
Non-Hydrogen Atoms Used in Refinement | |
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Non-Hydrogen Atoms | Number |
Protein Atoms | 2258 |
Nucleic Acid Atoms | |
Solvent Atoms | 243 |
Heterogen Atoms | 47 |
Software
Software | |
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Software Name | Purpose |
d*TREK | data scaling |
REFMAC | refinement |
PDB_EXTRACT | data extraction |
d*TREK | data reduction |
PHASER | phasing |