3NPX
Optimization of the in silico designed Kemp eliminase KE70 by computational design and directed evolution
X-RAY DIFFRACTION
Crystallization
| Crystalization Experiments | ||||
|---|---|---|---|---|
| ID | Method | pH | Temperature | Details |
| 1 | VAPOR DIFFUSION, SITTING DROP | 5.5 | 292 | 0.1M NaCl, 20% PEG 3000, pH 5.5, VAPOR DIFFUSION, SITTING DROP, temperature 292K |
| Crystal Properties | |
|---|---|
| Matthews coefficient | Solvent content |
| 2.51 | 50.95 |
Crystal Data
| Unit Cell | |
|---|---|
| Length ( Å ) | Angle ( ˚ ) |
| a = 52.135 | α = 90 |
| b = 83.331 | β = 90 |
| c = 129.203 | 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 | CCD | ADSC QUANTUM 315r | 2009-07-19 | M | SINGLE WAVELENGTH | ||||||
| Radiation Source | |||||
|---|---|---|---|---|---|
| ID # | Source | Type | Wavelength List | Synchrotron Site | Beamline |
| 1 | SYNCHROTRON | ESRF BEAMLINE ID14-4 | 0.975 | ESRF | ID14-4 |
Data Collection
| Overall | |||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| ID # | Resolution (High) | Resolution (Low) | Percent Possible (Observed) | R Merge I (Observed) | R Sym 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 | 1.79 | 50 | 99.6 | 0.091 | 0.095 | 28.7 | 9.5 | 53457 | 53243 | 19.03 | |||||||||
| Highest Resolution Shell | |||||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| ID # | Resolution (High) | Resolution (Low) | Percent Possible (All) | Percent Possible (Observed) | R Merge I (Observed) | R-Sym I (Observed) | Mean I Over Sigma (Observed) | Redundancy | Number Unique Reflections (All) | ||||||||||
| 1 | 1.8 | 1.86 | 99.1 | 0.31 | 0.29 | 5.9 | 8.4 | 5234 | |||||||||||
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 | 3NPU | 1.79 | 50 | 50467 | 2703 | 99.18 | 0.1983 | 0.19697 | 0.22313 | RANDOM | 19.029 | ||||||
| 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] | |
| 0.25 | -1.15 | 0.89 | ||||
| RMS Deviations | |
|---|---|
| Key | Refinement Restraint Deviation |
| r_dihedral_angle_2_deg | 30.214 |
| r_dihedral_angle_4_deg | 12.76 |
| r_dihedral_angle_3_deg | 11.691 |
| r_dihedral_angle_1_deg | 7.008 |
| r_scangle_it | 3.579 |
| r_scbond_it | 2.194 |
| r_angle_refined_deg | 1.258 |
| r_mcangle_it | 1.16 |
| r_mcbond_it | 0.712 |
| r_nbtor_refined | 0.298 |
| Non-Hydrogen Atoms Used in Refinement | |
|---|---|
| Non-Hydrogen Atoms | Number |
| Protein Atoms | 3698 |
| Nucleic Acid Atoms | |
| Solvent Atoms | 217 |
| Heterogen Atoms | |
Software
| Software | |
|---|---|
| Software Name | Purpose |
| HKL-2000 | data collection |
| PHASER | phasing |
| REFMAC | refinement |
| HKL-2000 | data reduction |
| SCALEPACK | data scaling |
