Generation of recombinant antibody fragments for membrane protein crystallization

Abstract

Membrane proteins are challenging targets for crystallization and structure determination by X-ray crystallography. Hurdles can be overcome by antibody-mediated crystallization. More than 25 unique structures of membrane protein:antibody complexes have already been determined. In the majority of cases, hybridoma-derived antibody fragments either in Fab or Fv fragment format were employed for these complexes. We will briefly introduce the background and current status of the strategy and describe in detail the current protocols of well-established methods for the immunization, the selection, and the characterization of antibodies, as well as the cloning, the production, and the purification of recombinant antibodies useful for structural analysis of membrane proteins.

Keywords: Antibodies; Crystallization; Fab fragment; Fv fragment; Membrane proteins; X-ray structures.