Proton transfer reactions on biosurfaces play an important role in a myriad of biological processes. Herein, the excited-state proton transfer reaction of 8-hydroxypyrene1,3,6-trisulfonate (HPTS) has been investigated in the presence of an important therapeutic protein, Protamine (PrS), using ground-state absorption, steady-state, and detailed time -resolved emission measurements. HPTS forms a 1:1 complex with Protamine with a high association constant of 2.6 X 10(4) M-1. The binding of HPTS with Protamine leads to a significant modulation in the ground-state prototropic equilibrium causing a downward shift of 1.1 unit in the acidity constant (pK(a)). In contrast to a large number of reports of slow proton transfer of HPTS on biosurfaces, interestingly, HPTS registers a faster proton transfer event in the presence of Protamine as compared to that of even the bulk aqueous buffer medium. Furthermore, the dimensionality of the proton diffusion process is also significantly reduced on the surface of Protamine that is in contrast to the behavior of HPTS in the bulk aqueous buffer medium, where the proton diffusion process is three-dimensional. The effect of ionic strength on the binding of HPTS toward PrS suggests a predominant role of electrostatic interaction between anionic HPTS and cationic Protamine, which is further supported by molecular docking simulations which predict that the most preferable binding site for HPTS on the surface of Protamine is surrounded by multiple cationic arginine residues.