Generalized Molecular Replacement

The following sections show how the 26-10 Fab fragment complexed with digoxin was solved by generalized molecular replacement (Brünger, 1991c). The space group of the 26-10 Fab/digoxin crystals is $P2_1$ (Strong, 1990), with the b-axis unique, and a non-crystallographic twofold symmetry is present. The following strategy was employed to solve the structure:
  1. Self-rotation function.
  2. Modification of the elbow angle of a known Fab structure.
  3. Cross-rotation function with the modified Fab structure.
  4. Filtering the rotation function by $PC$-refinement.
  5. Analysis of the $PC$-refinement.
  6. Translation function for molecule A, using the $PC$-refined model.
  7. Translation function for molecule B.
  8. Combined translation function to determine the relative position between A,B.
  9. Rigid-body refinement.
In general, one has to try several different starting elbow angles (spaced approximately 10$^{\circ}$ apart) and repeat steps 3-6 until a good solution is found. This strategy is also applicable to other multidomain proteins or PC-refinements of other degrees of freedom. The modification of the Fab example input files should be straightforward. An overview of the strategy is shown in Fig. 19.1.

Figure 19.1: Overview of molecular replacement.
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Xplor-NIH 2023-11-10