The antigen binding fragment of 10E8 (Fab) was prepared using LysC digestion, as previously described46 (link). The IgG was first reduced with 100 mM DTT for 1 h at 37 °C, followed by 1 h of dialysis in Hepes, pH 7.6, to reduce the DTT concentration to 1 mM. Antibodies were then dialyzed against 2 mM iodoacetamide for 48 h at 4 °C, and subjected to a final dialysis against Hepes, pH 7.6, for 2 h. After reduction and alkylation, antibodies were cleaved with Lys-C (Roche), run over a Protein A column to segregate away the Fc fragment, and then subjected to ion exchange (Mono S) and size-exclusion chromatography (S200). Purified 10E8 Fab was incubated with 10-fold excess peptide RRR-NEQELLELDKWASLWNWFDITNWLWYIR-RRR (American Peptide, CA) and the complex then set up set for 20°C vapor diffusion sitting drop crystallizations on the Honeybee 963 robot. 576 initial conditions adapted from the commercially available Hampton (Hampton Research), Precipitant Synergy (Emerald Biosystems), and Wizard (Emerald Biosystems) crystallization screens were set up and imaged using the Rockimager (Formulatrix), followed by hand optimization of crystal hits. Crystals were grown in 40% PEG 400, 0.1 M NaCitrate, 0.1 M Tris pH 7.5 diffracted to 2.1 Å resolution in a cryoprotectant composed of mother liquor supplemented with15% 2R-3R-butanediol and excess peptide. After mounting the crystals on a loop, they were flash cooled and data was collected at 1.00 Å wavelength at SER CAT ID-22 or BM-22 beamlines (APS) and processed using HKL-200047 . Structures were solved through molecular replacement with Phaser48 (link),49 (link), using a previously obtained free structure of 10E8 as a search model. Refinement of the structure was undertaken with Phenix50 (link), with iterative model building using Coot51 (link). The structure was validated with MolProbity52 (link), yielding 97% and 99.8% of residues falling within most favored Ramachandran regions and allowed Ramachandran regions, respectively. The structure was analyzed with APBS53 (link) for electrostatics, Ligplot54 (link) for direct contacts, PISA55 (link) for buried surface areas, and lsqkab (ccp4 Package56 (link)) for RMSD alignments. Helical wheels were generated using the program Pepwheel (http://150.185.138.86/cgi-bin/emboss/pepwheel). All graphics were prepared with Pymol (PyMOL Molecular Graphics System).