The recombinant protein Ultrabithorax (Ubx), a Drosophila melanogaster Hox transcription factor, self-assembles in vitro into biocompatible materials that are remarkably extensible and strong. Here, it is demonstrated that the strength of Ubx materials is due to intermolecular dityrosine bonds. Ubx materials autofluoresce blue, a characteristic of dityrosine, and bind dityrosine-specific antibodies. Monitoring the fluorescence of reduced Ubx fibers upon oxygen exposure reveals biphasic bond formation kinetics. Two dityrosine bonds in Ubx are identified by site-directed mutagenesis followed by measurements of fiber fluorescence intensity. One bond is located between the N-terminus and the homeodomain (Y4/Y296 or Y12/Y293), and another bond is formed by Y167 and Y240. Fiber fluorescence closely correlates with fiber strength, demonstrating that these bonds are intermolecular. This is the first identification of specific residues that participate in dityrosine bonds in protein-based materials. The percentage of Ubx molecules harboring both bonds can be decreased or increased by mutagenesis, providing an additional mechanism to control the mechanical properties of Ubx materials. Duplication of tyrosine-containing motifs in Ubx increases dityrosine content in Ubx fibers, suggesting these motifs could be inserted in other self-assembling proteins to strengthen the corresponding materials.