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 (Strong, 1990), with the
b-axis unique, and a non-crystallographic twofold symmetry is
present. The following strategy was employed to
solve the structure:
Self-rotation function.
Modification of the elbow angle of a known Fab structure.
Cross-rotation function with the modified Fab structure.
Filtering the rotation function by -refinement.
Analysis of the -refinement.
Translation function for molecule A, using the -refined model.
Translation function for molecule B.
Combined translation function to determine the relative position
between A,B.
Rigid-body refinement.
In general, one has to try several different starting elbow angles
(spaced approximately
10 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.