Optimized bacterial expression of a synthetic BRIL antibody.

Abstract

The use of monoclonal fragments antigen binding (Fabs) is a prevalent methodology facilitating protein structure determination via both crystallography and cryo-EM. The development of a synthetic Fab against the BRIL domain improved the accessibility of this approach, providing a general fiducial applicable to any protein of interest via the simple curation of a BRIL fusion protein. Here, we document the generation of a T7 Express ΔcybC strain allowing contaminant-free bacterial expression of the synthetic anti-BRIL Fab BAG2. We also report the crystal structure of BAG2 in complex with native cytochrome b₅₆₂, a complex arising from expression in canonical Escherichia coli strains.