Proton-Transfer Reaction Dynamics within the Human Serum Albumin Protein

Abstract

We report on femto- to nanosecond studies of the excited state intermolecular proton transfer (ESPT) reaction of trisodium 8-hydroxypyrene-1,3,6-trisulfonate (pyranine, HPTS) with the human serum albumin (HSA) protein. The formed robust 1:1 complexes (K(eq) = (2.6 ± 0.1) × 10(6) M(-1)) show both photoacid (∼430 nm) and conjugated photobase (∼500 nm) emissions of the caged HPTS in its protonated structure. The proton-transfer reactions in these complexes proceed in a large time window, spanning from 150 fs to ∼1.2 ns. The ultrafast component reflects a direct H-bond breaking and making in the robust complexes, involving the carboxylate groups of the amino acids, while the slowest one is arising from the slow dynamics of the so-called biological water. Additional time constants of the caged photoacid to give the conjugated photobase are observed, assigned to the ESPT reaction within "loose" complexes (3 to tens of picoseconds), and 130 ps and 1.2 ns due to the slow dynamics of the water molecules around the protein residues and involved in the proton transfer. The fs-ns anisotropy measurements confirm the robustness of the HPTS:HSA complexes. Our results indicate that, even though robust 1:1 complexes between HPTS and the HSA are formed, the system is heterogeneous, due to different possible interactions of the dye with the inside/outside parts of the protein. Furthermore, we find lower values of the initial anisotropy (r(0)) in the protein (0.33) and in γ-CD (0.28) in comparison with buffered aqueous solution (0.385). We propose that caging HPTS by the HSA protein and by the cyclodextrin affects the electronic redistribution in a different degree of mixing between the (1)L(a) and (1)L(b) states in the formed deprotonated form, for which the interactions of the sulfonate groups with the surroundings should play a key role.

Long‐Range Proton Conduction across Free‐Standing Serum Albumin Mats

“…The ability of HPTS to transfer protons to nearby amino acids has been observed before for the case of HPTS bound in the binding site of natively folded human serum albumin 38. It was also recently shown that HPTS can transfer protons to glucosamine units of chitosan 39.…”

Interaction of 1-pyrene sulfonic acid sodium salt with human serum albumin

“…Pyrene probes have strong absorption bands in the UV region, a high fluorescence quantum yield and long life time of the excited singlet state (can be examined with a nanosecond flash photolysis system). However, the literature concerning binding of pyrene and its derivatives to HSA or other serum albumins is not extensive [13,[19][20][21][22][23][24][25]. The albumin-induced changes of absorption and fluorescence spectra have been reported, but in most cases the authors do not specify unambiguously the probe/protein binding sites.…”

Drug/protein interactions studied by time-resolved fluorescence spectroscopy

“…[2][3][4][5][6]. These have shown the influence of the biological nanocavities not only on the diffusional/orientational motions of the drug, but also on its solvation dynamics and excited state processes such as energy, electron and proton transfer.…”