Role of predicted tannic acid deprotonation sites in Cu(II) coordination and human serum albumin interaction: Density functional theory and spectroscopic investigation.

The bioavailability of dietary polyphenols is critically dependent on their interaction with transport proteins like human serum albumin (HSA), yet how this process is modulated by metal ions remains poorly understood. This study elucidates the modulation of tannic acid (TA) binding to HSA by Cu(II) coordination at an atomic level. Integrating DFT calculations, multispectral techniques, and molecular simulations, this work characterizes the complete process: from TA deprotonation and subsequent Cu(II) coordination to its altered protein interaction. TA first undergoes sequential deprotonation at the 97-OH (pKa1 ≈ 6.09) and 98-OH (pKa2 ≈ 8.91) sites, existing predominantly as a monoanion at physiological pH (7.4). These anionic sites subsequently drive the formation of a stable, saturated 3:1 (Cu(II):TA) complex. Cu(II) coordination fundamentally altered TA's impact on HSA. While free TA quenched HSA's intrinsic fluorescence (via a mixed static-dynamic mechanism; Ksv ≈ 2.29 μM-1) and shortened its lifetime (from 1.17 ns to 0.72 ns), the Cu3TA complex exhibited reduced quenching via a purely static mechanism and extended the lifetime to 1.38 ns. Similarly, the complex substantially mitigated the profound secondary structure perturbations (α-helix change from 7.2 % to 3.6 %; β-sheet from -20.4 % to -11.2 %) caused by free TA. Molecular simulations revealed the TA-HSA binding mechanism is driven primarily by nonpolar interactions (van der Waals, hydrophobic effect) and is synergistically supported by electrostatics. A mechanism for this reduced affinity was confirmed: computational modeling of a simplified analogue, corroborated by experimental DLS and zeta potential data, pointed to two concurrent effects of Cu(II) coordination: steric hindrance (shielding key phenolic hydroxyls) and electrostatic neutralization. Collectively, this study establishes a mechanistic framework linking the polyphenol chemical state to transport protein interactions, offering critical insights into bioavailability.