METHODS
Protein Data Bank (.pdb) format files for poly leucine α-helices (28-mer), with either all trans or all gauche + rotamers, were generated using the BIOVIA Discovery Studio Visualizer software (Dassault Systèmes, San Diego). PDB files for GCN4 leucine zipper (1ZIK), 14-3-3 protein (2BQ0), Symfoil (3O4D) and Monofoil (3OL0) are from the RCSB database 15. All files were stripped of HETATOMs (i.e., waters and ligands). In the case of 14-3-3 the interface domain region relevant to the A domain was identified as residue positions A145-A206, and those for the B domain were identified as B3-B54/B98-B127 and these were the 3D printed regions (the B domains being monolithic). All residues for the GCN4 leucine zipper 30-mer were retained, as was the 42-mer region of the Monofoil polypeptide in the trimer oligomer forming a β-trefoil fold. Domain-swapped permutants of Monofoil 16 were generated using corresponding internal 42-mer regions of the Symfoil (i.e., intact β-trefoil) structure (Fig. 3).
Standard Tessellation Language (.stl) files were generated for all structures using the Chimera software package 17. Corresponding G-code files were generated using IdeaMaker software (Raise3D, Irvine CA) using standard van der Waals radii and a scale of 2.8 mm/Å. Molecules were 3D printed using a Raise3D Pro2 printer (Raise3D, Irvine CA) and 1.75 mm Flexmark 9 (Sigma Aldrich, St. Louis MO) thermoplastic polyurethane (TPU) filament (Durometer hardness of 90, elongation at break of 600%, and tensile strain of 55 MPa). Models were printed with 25% infill (grid pattern), three surface shells, and removable supports for regions of overhang. Printed models were stripped of supports, and weighed (with duplicate printed models typically exhibiting a mass difference within 1-2 g). Weight was used for quality control as printing nozzle obstruction yielded models of 5-10 g lower mass, weakened integrity, and reduced kinetic energy upon impact. An example of a 3D printed model, with supports trimmed, is given in Fig. 4.
Kinetic energy was imparted to models via an essentially elastic collision by fall from defined height onto a concrete surface. Ten repetitions were performed at each evaluated height, with essentially random model orientation. Drop heights varied from 5.0 cm to 12.0 m. At each height the number of repetitions resulting in model dissociation was noted and the fraction folded calculated. Drop heights were increased until the fraction folded yielded 0.0 (i.e., dissociation observed in all 10 trials) for each model. Kinetic energy was calculated as mass*g *height (g =9.8 m/s2) and the imparted energy (J) was normalized for total model mass (J/Kg).