Crystallization and preliminary X-ray analysis of a recombinant Fab fragment in complex with 17β-oestradiol

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Abstract
The recombinant Fab fragment of the anti-17beta-oestradiol antibody 57-2 has been a target for several protein-engineering experiments. A method for production, purification and crystallization of the Fab fragment alone (apo form) and in complex with the major female sex hormone 17beta-oestradiol is reported here. Diffracting apo-form crystals were only obtained with microseeding; crystals of the Fab-steroid complex were produced by co-crystallization in the presence of oestradiol and cross-seeding with the apo-form crystals. The crystals were grown using vapour-diffusion methods with reservoir solutions containing 10-14% PEG 4000 or 8-12% PEG 8000 and Tris-HCl buffer at high pH (9.0-9.5). Both the apo and complex crystals belong to space group P2(1)2(1)2(1) and diffract to 2.0 A resolution. High-resolution X-ray data sets suitable for structure determination were collected from flash-cooled crystals using 25% glycerol as the cryoprotectant.

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