2JZH
structure of IIB domain of the mannose transporter of E. coli
SOLUTION NMR
| NMR Experiment | ||||||||
|---|---|---|---|---|---|---|---|---|
| Experiment | Type | Sample Contents | Solvent | Ionic Strength | pH | Pressure | Temperature (K) | Spectrometer |
| 1 | 3D HCCH-TOCSY | 0.5-1 mM [U-100% 13C; U-100% 15N] protein, sodium phosphate, sodium azide | 100% D2O | 20 | 6.5 | ambient | 303 | |
| 2 | 3D HCCH-COSY | 0.5-1 mM [U-100% 13C; U-100% 15N] protein, sodium phosphate, sodium azide | 100% D2O | 20 | 6.5 | ambient | 303 | |
| 3 | 3D 1H-13C NOESY | 0.5-1 mM [U-100% 13C; U-100% 15N] protein, sodium phosphate, sodium azide | 100% D2O | 20 | 6.5 | ambient | 303 | |
| 4 | 2D 1H-15N HSQC | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 5 | 3D 1H-15N NOESY | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 6 | 3D CBCA(CO)NH | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 7 | 3D C(CO)NH | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 8 | 3D HNCACB | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 9 | 3D HBHA(CO)NH | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| 10 | 3D H(CCO)NH | 0.5-1 mM [U-100% 15N] protein, sodium phosphate, sodium azide | 90% H2O/10% D2O | 20 | 6.5 | ambient | 303 | |
| NMR Spectrometer Information | |||
|---|---|---|---|
| Spectrometer | Manufacturer | Model | Field Strength |
| 1 | Bruker | DRX | 600 |
| 2 | Bruker | DRX | 800 |
| 3 | Bruker | DRX | 500 |
| NMR Refinement | ||
|---|---|---|
| Method | Details | Software |
| simulated annealing in torsion angle space | RMS DEVIATION FROM EXPERIMENTAL RESTRAINTS (NUNMER OF RESTRAINTS IN PARENTHESES): INTERPROTON DISTANCES (A) (1577) 0.008, TORSION ANGLES (DEG) (478) 0.26, 13CALPHA SHIFTS (PPM) (161) 1.25, 13CBETA SHIFTS (PPM) (158) 1.23. RDC R-factors (%): PHAGE 1DNH (151) 4.2, PHAGE 1DNC' (113) 18.2, PHAGE 2DHNC' (113) 16.5, PEG/HEXANOL 1DNH (141) 6.0, PEG/HEXANOL 1DNC' (96) 25.8, PEG/HEXANOL 2DHNC' (103) 23.5. | PIPP |
| NMR Ensemble Information | |
|---|---|
| Conformer Selection Criteria | all calculated structures |
| Conformers Calculated Total Number | 130 |
| Conformers Submitted Total Number | 1 |
| Representative Model | 1 (minimized average structure) |
| Additional NMR Experimental Information | |
|---|---|
| Details | Double and triple resonance 3D NMR experiments for assignments (HNCACB, CBCA(CO)NH, HBHA(CBCACO)NH, C(CCO)NH, H(CCO)NH, HCCH-COSY, HCCH-TOCSY). NOE-derived interproton distance restrants from 3D 15N-, 13C-, 13C/15M-, 13C/13C, and 15N/15N-separated NOE spectra. Side chain rotamers from 3H heteronuclear couplings and short mixing time 3D 13C-separated NOE and 3D 15N-seaparated ROE spectra. RDCs obtained by taking difference in J couplings in aligned (phage pf1 and PEG/hexanol) and isotopic media. |
| Computation: NMR Software | ||||
|---|---|---|---|---|
| # | Classification | Version | Software Name | Author |
| 1 | chemical shift assignment | PIPP | Garrett and Clore | |
| 2 | data analysis | PIPP | Garrett and Clore | |
| 3 | refinement | X-PLOR NIH | 2.18.1 | Schwieters, Kuszewski and Clore |
| 4 | chemical shift assignment | CAPP | Garrett and Clore | |
| 5 | processing | NMRPipe | Delaglio, Grzesiek, Vuister, Zhu, Pfeifer and Bax | |
