Evaluation of structure and morphology of extruded wheat gluten (WG) films showed WG protein assemblies elucidated on a range of length scales from nano (4.4 angstrom and 9 to 10 angstrom, up to 70 angstrom) to micro (10 mu m). The presence of NaOH in WG films induced a tetragonal structure with unit cell parameters, a = 51.85 angstrom and c = 40.65 angstrom, whereas NH4OH resulted in a bidimensional hexagonal close-packed (HCP) structure with a lattice parameter of 70 angstrom. In the WG films with NH4OH, a highly polymerized protein pattern with intimately mixed glutenins and gliadins bounded through SH/SS interchange reactions was found. A large content of beta-sheet structures was also found in these films, and the film structure was oriented in the extrusion direction. In conclusion, this study highlights complexities of the supramolecular structures and conformations of wheat gluten polymeric proteins in biofilms not previously reported for biobased materials.