2VEM
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
Starting Model(s)
| Initial Refinement Model(s) | |||
|---|---|---|---|
| Type | Source | Accession Code | Details |
| experimental model | PDB | 1DKW | PDB ENTRY 1DKW |
Crystallization
| Crystalization Experiments | ||||
|---|---|---|---|---|
| ID | Method | pH | Temperature | Details |
| 1 | 5.5 | 20% PEG6000, 2,5% T-BUTANOL, 0.1 M CITRIC ACID PH 5,5 | ||
| Crystal Properties | |
|---|---|
| Matthews coefficient | Solvent content |
| 2.23 | 44.8 |
Crystal Data
| Unit Cell | |
|---|---|
| Length ( Å ) | Angle ( ˚ ) |
| a = 45.8 | α = 90 |
| b = 86.9 | β = 97.2 |
| c = 56.4 | 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 | M | SINGLE WAVELENGTH | |||||||
| Radiation Source | |||||
|---|---|---|---|---|---|
| ID # | Source | Type | Wavelength List | Synchrotron Site | Beamline |
| 1 | SYNCHROTRON | EMBL/DESY, HAMBURG BEAMLINE X13 | EMBL/DESY, HAMBURG | X13 | |
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 | 2.2 | 25 | 99.8 | 0.14 | 14.36 | 7.6 | 22262 | 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 | 2.2 | 2.3 | 100 | 0.62 | 3.63 | 7.6 | |||||||||||||
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 | 2.2 | 19.66 | 21148 | 1114 | 100 | 0.196 | 0.193 | 0.247 | RANDOM | 22.51 | ||||||
| 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] | |
| -0.17 | 0.15 | 0.47 | -0.26 | |||
| RMS Deviations | |
|---|---|
| Key | Refinement Restraint Deviation |
| r_dihedral_angle_2_deg | 35.778 |
| r_dihedral_angle_3_deg | 17.857 |
| r_dihedral_angle_4_deg | 12.813 |
| r_dihedral_angle_1_deg | 6.303 |
| r_scangle_it | 2.585 |
| r_scbond_it | 1.646 |
| r_angle_refined_deg | 1.405 |
| r_mcangle_it | 1.103 |
| r_mcbond_it | 0.65 |
| r_nbtor_refined | 0.3 |
| Non-Hydrogen Atoms Used in Refinement | |
|---|---|
| Non-Hydrogen Atoms | Number |
| Protein Atoms | 3593 |
| Nucleic Acid Atoms | |
| Solvent Atoms | 98 |
| Heterogen Atoms | 23 |
Software
| Software | |
|---|---|
| Software Name | Purpose |
| REFMAC | refinement |
| XDS | data reduction |
| XDS | data scaling |
| MOLREP | phasing |
