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| F is for Fab Fragments |
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| Fab and Fv fragments can be used as co-crystallization agents. Fab fragments have decent solubility properties and bind specifically to antigens with reasonable equilibrium constants (105 to 108 M-1). Fab fragments can, in some instances, effectively transform aggregated protein into soluble, monodisperse protein, suitable for crystallization trials. Since antibodies are frequently available from related biochemical studies, they can often be applied more readily than molecular biology (point mutants, truncations, molecular engineering). However, it seems MOBO gets faster and easier with each passing year. It is often useful to consider screening several different Fab fragments that recognize different epitopes for crystallization trials. Sometimes Fab fragments can immobilize a region of a protein sample, reducing sample flexibility, enhancing conformational homogeneity of the sample which in turn can enhance chances for crystallization.
Using an Fv fragment in lieu of a Fab fragment might have some advantage since there is no flexible elbow to inhibit crystallization.
When preparing antigen-antibody complexes for crystallization, one should carefully select the antibody, prepare a homogeneous Fab species, and prepare the Fab-sample complex with proper stoichiometry. Ideally the Fab should not interfere with the native sample conformation or activity.
The presence of a Fab fragment in the crystal can assist in the crystallographic structure determination by allowing one to utilize molecular replacement.
Fab and Fv fragments should be considered useful tools in the crystallization toolbox for manipulating sample solubility, conformational flexibility, and crystal lattice contacts, as well as being a possible tool for molecular replacement.
Reference:
The use of antibody fragments for crystallization and structure determinations. LC Kovari, C. Momany and MG Rossman. Structure 15 December 1995, 3: 1291-1293. |
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