2OEU
Full-length hammerhead ribozyme with Mn(II) bound
Experimental procedure
| Experimental method | SINGLE WAVELENGTH |
| Source type | SYNCHROTRON |
| Source details | SSRL BEAMLINE BL9-1 |
| Synchrotron site | SSRL |
| Beamline | BL9-1 |
| Temperature [K] | 100 |
| Detector technology | CCD |
| Collection date | 2006-03-15 |
| Detector | ADSC QUANTUM 4 |
| Wavelength(s) | 1.37756 |
| Spacegroup name | C 1 2 1 |
| Unit cell lengths | 50.312, 68.981, 60.443 |
| Unit cell angles | 90.00, 112.62, 90.00 |
Refinement procedure
| Resolution | 55.800 - 2.000 |
| R-factor | 0.17143 |
| Rwork | 0.166 |
| R-free | 0.22087 |
| Structure solution method | same cell as 2goz |
| Starting model (for MR) | 2goz |
| RMSD bond length | 0.010 |
| RMSD bond angle | 1.728 |
| Data reduction software | MOSFLM |
| Data scaling software | CCP4 ((SCALA)) |
| Refinement software | REFMAC (5.3.0021) |
Data quality characteristics
| Overall | Outer shell | |
| Low resolution limit [Å] | 38.520 | 2.110 |
| High resolution limit [Å] | 2.000 | 2.000 |
| Rmerge | 0.037 | 0.113 |
| Number of reflections | 12527 | |
| <I/σ(I)> | 23.2 | 10.4 |
| Completeness [%] | 97.0 | 96.3 |
| Redundancy | 4.2 | 4.2 |
Crystallization Conditions
| crystal ID | method | pH | temperature | details |
| 1 | VAPOR DIFFUSION, HANGING DROP | 6.5 | 301 | THE ENZYME AND SUBSTRATE WERE MIXED IN EQUIMOLAR AMOUNTS IN A SOLUTION CONTAINING 50 MM MES PH 5.5, 1.5 MM EDTA. THE COMPLEX WAS FORMED BY INCUBATING THE MIXTURE AT 95C FOR 2 MIN., THEN AT 65C FOR 2 MIN., AND FINALLY AT 27C FOR 5 MIN. 1 MM MGCL2 WAS INCLUDED IN THE MIXTURE BEFORE THE FINAL INCUBATION STEP. A 10 MG/ML CONCENTRATION OF RNA WAS USED FOR THE CRYSTALLIZATION EXPERIMENTS. THE RESERVOIR SOLUTION CONTAINED 0.5M (NH4)2SO4, 100 MM MES PH 6.5, AND 35% PEG 3350. AFTER MIXING THE RESERVOIR SOLUTION, THE SALT AND PEG PHASES WERE ALLOWED TO SEPARATE, AND ONLY THE SALT PHASE WAS USED FOR THE DROPS. THE CRYSTALS GREW IN HANGING DROPS OF 2 MICROLITERS AFTER 12 MONTHS OF INCUBATION AT 28C. THE CRYSTALS WERE WASHED IN THE SALT PHASE OF THE RESERVOIR SOLUTION PRIOR TO CRYO-FREEZING, VAPOR DIFFUSION, HANGING DROP, temperature 301K |
Crystallization Reagents
| ID | crystal ID | solution ID | reagent name | concentration | details |
| 1 | 1 | 1 | MES | ||
| 10 | 1 | 2 | PEG | ||
| 11 | 1 | 2 | (NH4)2SO4 | ||
| 2 | 1 | 1 | EDTA | ||
| 3 | 1 | 1 | MGCL2 | ||
| 4 | 1 | 1 | PEG | ||
| 5 | 1 | 1 | (NH4)2SO4 | ||
| 6 | 1 | 1 | H2O | ||
| 7 | 1 | 1 | MES | ||
| 8 | 1 | 2 | EDTA | ||
| 9 | 1 | 2 | MGCL2 |
