2MA4
Solution NMR Structure of yahO protein from Salmonella typhimurium, Northeast Structural Genomics Consortium (NESG) Target StR106
SOLUTION NMR
| NMR Experiment | ||||||||
|---|---|---|---|---|---|---|---|---|
| Experiment | Type | Sample Contents | Solvent | Ionic Strength | pH | Pressure | Temperature (K) | Spectrometer |
| 1 | 2D 1H-15N HSQC | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 2 | 2D 1H-13C HSQC aliphatic | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 3 | 3D HNCO | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 4 | (4,3)D GFT CABCA(CO)NHN | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 5 | (4,3)D GFT HNHNCABCA | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 6 | (4,3)D GFT HABCAB(CO)NHN | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 7 | 3D 1H-15N,13C NOESY | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 8 | (4,3)D GFT HCCH-COSY aliphatic | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 9 | 3D (H)CCH-COSY aromatic | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 10 | 3D (H)CCH-TOCSY aliphatic | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 11 | 2D 1H-13C HSQC aromatic | 1.1 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 12 | 2D 1H-13C HSQC methyl | 1 mM [5% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 90% H2O/10% D2O | 4.5 | ambient | 298 | ||
| 13 | 2D 1H-15N HSQC | 0.3 mM [U-100% 13C; U-100% 15N] StR106, 0.02 % NaN3, 10 mM DTT, 5 mM calcium chloride, 100 mM sodium chloride, 20 mM ammonium acetate, 50 uM DSS | 100% D2O | 4.5 | ambient | 298 | ||
| NMR Spectrometer Information | |||
|---|---|---|---|
| Spectrometer | Manufacturer | Model | Field Strength |
| 2 | Varian | INOVA | 750 |
| NMR Refinement | ||
|---|---|---|
| Method | Details | Software |
| simulated annealing | STRUCTURE DETERMINATION WAS PERFORMED BY RUNNING CYANA AND ASDP IN PARALLEL USING NOE-BASED CONSTRAINTS AND PHI AND PSI DIHEDRAL ANGLE CONSTRAINTS FROM TALOS+. CONSENSUS PEAK ASSIGNMENTS WERE SELECTED AND USED IN ITERATIVE REFINEMENT WITH CYANA. THE 20 CONFORMERS OUT OF 100 WITH THE LOWEST TARGET FUNCTION WERE FURTHER REFINED BY SIMULATED ANNEALING IN EXPLICIT WATER BATH USING THE PROGRAM CNS WITH PARAM19 FORCE FIELD | CNS |
| NMR Ensemble Information | |
|---|---|
| Conformer Selection Criteria | target function |
| Conformers Calculated Total Number | 100 |
| Conformers Submitted Total Number | 20 |
| Representative Model | 1 (lowest energy) |
| Computation: NMR Software | ||||
|---|---|---|---|---|
| # | Classification | Version | Software Name | Author |
| 1 | refinement | CNS | 1.3 | Brunger, Adams, Clore, Gros, Nilges and Read |
| 2 | refinement | CYANA | 3.0 | Guntert, Mumenthaler and Wuthrich |
| 3 | geometry optimization | CYANA | 3.0 | Guntert, Mumenthaler and Wuthrich |
| 4 | structure solution | CYANA | 3.0 | Guntert, Mumenthaler and Wuthrich |
| 5 | structure calculation | AS-DP | 1.0 | Huang, Tejero, Powers and Montelione |
| 6 | refinement | AS-DP | 1.0 | Huang, Tejero, Powers and Montelione |
| 7 | data analysis | XEASY | 1.3.13 | Bartels et al. |
| 8 | peak picking | XEASY | 1.3.13 | Bartels et al. |
| 9 | chemical shift assignment | XEASY | 1.3.13 | Bartels et al. |
| 10 | data analysis | XEASY | 1.3.13 | Bartels et al. |
| 11 | data analysis | XEASY | 1.3.13 | Keller and Wuthrich |
| 12 | peak picking | XEASY | 1.3.13 | Keller and Wuthrich |
| 13 | chemical shift assignment | XEASY | 1.3.13 | Keller and Wuthrich |
| 14 | data analysis | XEASY | 1.3.13 | Keller and Wuthrich |
| 15 | collection | VnmrJ | 2.2D | Varian |
| 16 | geometry optimization | TALOS+ | Shen, Cornilescu, Delaglio and Bax | |
| 17 | structure validation | PSVS | 1.4 | Bhattacharya, Montelione |
| 18 | data analysis,peak picking,chemical shift assignment | XEASY | 1.3.13 | Bartels et al. |
| 19 | data analysis | XEASY | 1.3.13 | Bartels et al. |
| 20 | data analysis,peak picking,chemical shift assignment | XEASY | 1.3.13 | Keller and Wuthrich |
| 21 | data analysis | XEASY | 1.3.13 | Keller and Wuthrich |
| 22 | processing | NMRPipe | 2.3 | Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax |
| 23 | processing | PROSA | 6.4 | Guntert |
| 24 | data analysis | DYANA | 1.5 | Guntert, Braun and Wuthrich |
| 25 | data analysis | UBNMR | 1.0 | Shen |
