The water-soluble porphyrin meso-tetrakis(p-sulfonatophenyl)porphyrin (TSPP) can be noncovalently bound to tubulin and used as a photosensitizer, which upon irradiation triggers photochemical reactions that lead to conformational changes of the protein. These conformational changes in turn inhibit tubulin's primary function of polymerizing into microtubules. We explored the possibility of using two-photon excitation of the bound porphyrin to induce photosensitized protein unfolding. Although TSPP has a relatively low cross section (similar to 30 GM) our results did find that two-photon excitation of the ligand causes partial unfolding of the tubulin host and the inhibition of the in vitro formation of microtubules. Conversely, irradiating tubulin alone caused no such effects despite the large irradiance per pulse (97-190 GW/cm(2)). The conformational changes were characterized using spectroscopic studies and provide a promising protocol for the future application of non-native photosensitization of proteins.