5JC7
Crystal structure of chicken MDA5 with 5'p 24-mer dsRNA and ADP-Mg2+ at 2.75 A resolution.
X-RAY DIFFRACTION
Crystallization
Crystalization Experiments | ||||
---|---|---|---|---|
ID | Method | pH | Temperature | Details |
1 | VAPOR DIFFUSION, SITTING DROP | 7.5 | 293 | Directly after size exclusion chromatography chMDA5 deltaCARD-Q was mixed with 24-mer dsRNA in a 0.5:1 molar ratio and incubated for 30 minutes on ice. The complexes were concentrated to around 10 mg/ml and 2 mM AMPPNP and 200 mM NDSB211 (Dimethyl(2-hydroxyethyl)ammonium propane sulfonate) were added. chMDA5 delatCARD-Q:24bp dsRNA:AMPPNP was mixed with reservoir buffer (0.1 mM Hepes pH 7.5, 11-12% PEG 3350, 2-4% sucrose) in a 2:1 ratio. Crystals grew in three days at 20 degree and were harvested in cryo-protecting solution (0.1 mM Hepes pH 7.5, 25% 3350, 10% ethylene glycol) before flash freezing with liquid nitrogen |
Crystal Properties | |
---|---|
Matthews coefficient | Solvent content |
2.57 | 52.18 |
Crystal Data
Unit Cell | |
---|---|
Length ( Å ) | Angle ( ˚ ) |
a = 99.75 | α = 90 |
b = 133.4 | β = 90 |
c = 138.44 | γ = 90 |
Symmetry | |
---|---|
Space Group | P 21 21 21 |
Diffraction
Diffraction Experiment | ||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
ID # | Crystal ID | Scattering Type | Data Collection Temperature | Detector | Detector Type | Details | Collection Date | Monochromator | Protocol | |||||
1 | 1 | x-ray | 100 | PIXEL | DECTRIS PILATUS3 6M | 2013-07-25 | M | SINGLE WAVELENGTH |
Radiation Source | |||||
---|---|---|---|---|---|
ID # | Source | Type | Wavelength List | Synchrotron Site | Beamline |
1 | SYNCHROTRON | ESRF BEAMLINE ID29 | 0.9724 | ESRF | ID29 |
Data Collection
Overall | |||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
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 | 2.75 | 50 | 99.1 | 0.168 | 6.58 | 4.66 | 48273 |
Highest Resolution Shell | |||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
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 | 2.75 | 2.82 | 99.2 | 0.809 | 1.52 | 4.89 |
Refinement
Statistics | |||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
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 | R-Free | R-Free Selection Details | Mean Isotropic B | ||||||
X-RAY DIFFRACTION | MOLECULAR REPLACEMENT | THROUGHOUT | 5JC3 | 2.75 | 50 | 45840 | 2433 | 99.08 | 0.29224 | 0.29064 | 0.32199 | RANDOM | 69.662 |
Temperature Factor Modeling | ||||||
---|---|---|---|---|---|---|
Anisotropic B[1][1] | Anisotropic B[1][2] | Anisotropic B[1][3] | Anisotropic B[2][2] | Anisotropic B[2][3] | Anisotropic B[3][3] | |
7.4 | 2.21 | -9.61 |
RMS Deviations | |
---|---|
Key | Refinement Restraint Deviation |
r_dihedral_angle_2_deg | 36.299 |
r_dihedral_angle_3_deg | 16.126 |
r_dihedral_angle_4_deg | 14.386 |
r_long_range_B_refined | 7.322 |
r_long_range_B_other | 7.322 |
r_dihedral_angle_1_deg | 6.279 |
r_mcangle_it | 5.072 |
r_mcangle_other | 5.072 |
r_scangle_other | 4.224 |
r_mcbond_it | 2.972 |
Non-Hydrogen Atoms Used in Refinement | |
---|---|
Non-Hydrogen Atoms | Number |
Protein Atoms | 10537 |
Nucleic Acid Atoms | 1021 |
Solvent Atoms | |
Heterogen Atoms | 58 |
Software
Software | |
---|---|
Software Name | Purpose |
REFMAC | refinement |
XDS | data reduction |
XSCALE | data scaling |
PHASER | phasing |