Micro-Crystallography
Crystals of N2C/D282C rhodopsin had the form of small micro-crystalline needles measuring ~5x5x90 µm3. Due to this small size the micro-crystallographic setup at ID13, ESRF, Grenoble [5, 6] was necessary to collect data of sufficient quality for structure determination. The achieved diffraction limit was better than 3.5 Å in ~10% of the crystals, a ten times higher success rate than previously achieved with crystals grown from native rhodopsin [7].
However, the high dose delivered to the small crystal volume reduced the effective resolution to less then 5 Å after 8 frames had been collected. To minimize the effect of radiation damage, data was therefore collected from 15 positions along a single crystalline needle. A consistent and complete dataset was obtained by merging data from the best 10 positions.
Data collection and refinement of N2C/D282C rhodopsin |
|
| Data collection | |
| Space group | P31 |
| Cell dimensions | |
| a, b, c (Å) | 109.3, 109.3, 77.7 |
| α, β, γ (°) | 90, 90, 120 |
| Resolution (Å) | 50-3.4 (3.5-3.4) |
| Rmerge | 0.24 (0.703) |
| I / sI | 5.42 (1.79) |
| Completeness (%) | 95.9 (92.3) |
| Redundancy | 3.0 (2.6) |
| Molecular Replacement with Monomer A of 1GZM | |
| R-factor | 0.455 |
| Correlation coefficient | 0.344 |
| Refinement | |
| Resolution (Å) | 50-3.4 |
| No. reflections | 13282 |
| Rwork / Rfree | 0.287/0.325 |
| No. atoms | |
| Protein | 5178 |
| Ligand/ion | 67 |
| R.m.s deviations | |
| Bond lengths (Å) | 0.014 |
| Bond angles (°) | 1.66 |
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