2VEI
Structure-based enzyme engineering efforts with an inactive monomeric TIM variant: the importance of a single point mutation for generating an active site with suitable binding properties
X-RAY DIFFRACTION
Crystallization
| Crystalization Experiments | ||||
|---|---|---|---|---|
| ID | Method | pH | Temperature | Details |
| 1 | 8.5 | 0.1 M TRIS/HCL PH 8.5, 1.9 M MGSO4 | ||
| Crystal Properties | |
|---|---|
| Matthews coefficient | Solvent content |
| 2.58 | 52.42 |
Crystal Data
| Unit Cell | |
|---|---|
| Length ( Å ) | Angle ( ˚ ) |
| a = 91.24 | α = 90 |
| b = 52.59 | β = 119.04 |
| c = 92.38 | γ = 90 |
| Symmetry | |
|---|---|
| Space Group | P 1 21 1 |
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 | MARRESEARCH | PROPHYSICS XRM-216 | 2001-05-13 | M | SINGLE WAVELENGTH | |||||
| Radiation Source | |||||
|---|---|---|---|---|---|
| ID # | Source | Type | Wavelength List | Synchrotron Site | Beamline |
| 1 | ROTATING ANODE | ENRAF-NONIUS FR591 | |||
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 | 1.89 | 25 | 99.3 | 0.06 | 18.4 | 3.8 | 61427 | 3 | |||||||||||
| 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 | 1.89 | 1.95 | 99.7 | 0.15 | 7.32 | 3.7 | |||||||||||||
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 | PDB ENTRY 1DKW | 1.89 | 18.85 | 58355 | 3072 | 100 | 0.168 | 0.166 | 0.215 | RANDOM | 12.99 | ||||||
| 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] | |
| -0.22 | 0.1 | 0.35 | -0.03 | |||
| RMS Deviations | |
|---|---|
| Key | Refinement Restraint Deviation |
| r_dihedral_angle_2_deg | 34.632 |
| r_dihedral_angle_4_deg | 16.473 |
| r_dihedral_angle_3_deg | 13.493 |
| r_dihedral_angle_1_deg | 5.85 |
| r_scangle_it | 3.168 |
| r_scbond_it | 2.029 |
| r_mcangle_it | 1.203 |
| r_mcbond_it | 0.73 |
| r_nbtor_refined | 0.303 |
| r_symmetry_hbond_refined | 0.284 |
| Non-Hydrogen Atoms Used in Refinement | |
|---|---|
| Non-Hydrogen Atoms | Number |
| Protein Atoms | 5433 |
| Nucleic Acid Atoms | |
| Solvent Atoms | 776 |
| Heterogen Atoms | 80 |
Software
| Software | |
|---|---|
| Software Name | Purpose |
| REFMAC | refinement |
| XDS | data reduction |
| XDS | data scaling |
| MOLREP | phasing |
