Bioluminescence Resonance Energy Transfer (BRET)-Mediated Protein Release from Self-Illuminating Photoresponsive Biomaterials

Phototriggered release of various cargos, including soluble protein factors and small molecules, has the potential to correct aberrant biological events by offering spatiotemporal control over local therapeutic levels. However, the poor penetration depth of light historically limits implementation to subdermal regions, necessitating alternative methods of light delivery to achieve the full potential of photodynamic therapeutic release. Here, we introduce a strategy exploiting bioluminescence resonance energy transfer (BRET)–an energy transfer process between light-emitting Nanoluciferase (NLuc) and a photosensitive acceptor molecule–to drive biomolecule release from hydrogel biomaterials. Through a facile, one-pot, and high-yielding synthesis (60–70%), we synthesized a heterobifunctional ruthenium cross-linker bearing an aldehyde and an azide (CHO-Ru-N₃), a compound that we demonstrate undergoes predictable exchange of the azide-bearing ligand under blue-green light irradiation (>550 nm). Following site-specific conjugation to NLuc via sortase-tag enhanced protein ligation (STEPL), the modified protein was covalently attached to a poly(ethylene glycol) (PEG)-based hydrogel via strain-promoted azide–alkyne cycloaddition (SPAAC). Leveraging the high photosensitivity of Ru compounds, we demonstrate rapid and equivalent release of epidermal growth factor (EGF) via either direct illumination or via BRET-based bioluminolysis. As NLuc-originated luminescence can be controlled equivalently throughout the body, we anticipate that this unique protein release strategy will find use for locally triggered drug delivery following systemic administration of a small molecule.