Modulation of prototropic activity and rotational relaxation dynamics of a cationic biological photosensitizer within the motionally constrained bio-environment of a protein.

The present work describes the interaction of a promising cancer cell photosensitizer, harmane (HM), with a model transport protein, Bovine Serum Albumin (BSA). The studied molecule of interest (HM) belongs to the family of naturally occurring fluorescent drug-binding alkaloids, the β-carbolines. A combined use of steady-state and time-resolved fluorescence techniques is applied to follow and characterize the binding interaction. The polarity-dependent prototropic activity of HM is found to be responsible for the commendable sensitivity of the probe to the protein environments and is distinctly reflected on the emission profile. Steady-state fluorescence anisotropy study reveals the impartation of a considerable degree of motional restriction on the drug molecule as a result of binding to the protein. Contrary to the single-exponential nature of fluorescence anisotropy decay of HM in aqueous buffer, they are found to be biexponential in the protein environment. The rotational relaxation dynamics of HM within the protein has been interpreted on the lexicon of the Two-Step and Wobbling-in-Cone model. The probable binding location for the cationic drug is found to be the hydrophilic binding zone of BSA, i.e., domain I (characterized by a net negative charge). The AutoDock-based blind docking simulation has been explored for evaluating an unbiased result of the probable interaction site of HM in the protein. To unfold the effect of binding of the drug on the secondary structural content of the protein, circular dichroism (CD) spectroscopy has been exploited to see that binding of the drug accompanies some decrease in α-helical content of BSA, and the effect gradually saturates toward a higher drug/protein molar ratio.