Photoinduced unfolding of tubulin dimers bound to meso-tetrakis (sulfonatophenyl) porphyrin

Partial Unfolding of Tubulin Heterodimers Induced by Two-Photon Excitation of Bound meso-Tetrakis(sulfonatophenyl)porphyrin

“…Effects of TPE Irradiation on the OPE Emission of Tubulin. In previous experiments, the OPE irradiation of the complex revealed that photosensitization by the porphyrin ligand produced a large decrease of the intrinsic emission of tubulin, consistent with the occurrence of conformational modifications of the protein 12 (while there appeared to be no depletion of the ground state due to permanent chemical changes of the Trp residues). In the case of TPE, a similar effect was observed.…”

“…In both cases an additional 1 nm redshift of the Soret band is observed after irradiation of the TSPP/tubulin complex (Figure 1, parts A and B) and is in agreement with what was observed with OPE. 12 Thus, in both cases (free and bound TSPP) the 80 MHz laser produces photobleaching while the 1 kHz does not despite having comparable photon density at the sample. How can this difference be reconciled?…”

“…The absence of an effect in free TSPP is consistent with what is shown by the absorption (Figure 1), but is in contrast to previous evidence that OPE produces significant bleaching of free TSPP. 12 As explained earlier, this difference could be caused by the fact that TPE prompts a transition to a molecular orbital distinct from the one occupied upon OPE because of different symmetry selection rules from the ground state. In such case, the photochemical processes that lead to bleaching upon OPE might be reduced or prevented if, for instance, the excited state populated by TPE has a different relaxation pathway than the one populated by OPE.…”

“…10,11 A similar approach was taken to demonstrate that meso-tetrakis(p-sulfonatophenyl)porphyrin (TSPP) is able to prompt conformational and functional changes on larger proteins, i.e., tubulin, through one-photon excitation (OPE) at low irradiance. 12 Tubulin is a globular protein which assembles into heterodimers that form the basic building component for microtubules (MT). 13,14 MTs are ubiquitous in eukaryotic cells and perform important functions such as intracellular transport, morphogenesis, mitosis, and meiosis.…”

“…Having demonstrated the ability to produce photosensitized inhibition of MT through OPE, 12 we have investigated whether similar effects are produced upon two-photon excitation (TPE) of the porphyrin using NIR excitation at 805−810 nm (corresponding to the OPE at ∼402.5−405 nm). TPE involves the near simultaneous absorption of two photons in which the first photon excites the molecule to a virtual state.…”